hin diii  (New England Biolabs)


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    HindIII
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    HindIII 50 000 units
    Catalog Number:
    r0104l
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    249
    Size:
    50 000 units
    Category:
    Restriction Enzymes
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    Structured Review

    New England Biolabs hin diii
    HindIII
    HindIII 50 000 units
    https://www.bioz.com/result/hin diii/product/New England Biolabs
    Average 99 stars, based on 27 article reviews
    Price from $9.99 to $1999.99
    hin diii - by Bioz Stars, 2020-08
    99/100 stars

    Images

    1) Product Images from "Local chromosome context is a major determinant of crossover pathway biochemistry during budding yeast meiosis"

    Article Title: Local chromosome context is a major determinant of crossover pathway biochemistry during budding yeast meiosis

    Journal: eLife

    doi: 10.7554/eLife.19669

    Southern blots of Hin dIII and Hin dIII-VDE digests of DNA from spo11 strains with inserts at HIS4 (top) and at URA3 (bottom). Gel labels are as in Figure 1 ; JM—joint molecule recombination intermediates. DOI: http://dx.doi.org/10.7554/eLife.19669.015
    Figure Legend Snippet: Southern blots of Hin dIII and Hin dIII-VDE digests of DNA from spo11 strains with inserts at HIS4 (top) and at URA3 (bottom). Gel labels are as in Figure 1 ; JM—joint molecule recombination intermediates. DOI: http://dx.doi.org/10.7554/eLife.19669.015

    Techniques Used:

    Spo11-initiated events at the two insert loci. ( A ) Spo11-catalyzed DSBs are more frequent at HIS4 that at URA3 . Left—Southern blots of Eco RI digests of DNA from vde∆ strains, probed with pBR322 sequences, showing Spo11-DSBs in the Parent 2 insert (see Figure 1 ) in resection/repair-deficient sae2∆ mutant strains. Right—location of DSBs and probe and DSB frequencies (average of 7 and 8 hr samples from a single experiment; error bars represent range). Spo11-DSBs in the Parent 1 inserts at HIS4 and URA3 were at different locations within the insert, but displayed similar ratios between the two loci (data not shown). ( B ) Southern blots of Hin dIII digests of DNA from vde∆ strains, to detect total Spo11-initiated crossovers. ( C ) Southern blots of Hin dIII-VDE double digests of the same samples, to determine the background contribution of Spo11-initiated COs in subsequent experiments measuring VDE-initiated COs, which will be VDE-resistant due to conversion of the VRS site to VRS103 . Probes were as shown in Figure 1 . ( D ) Quantification of data in panels B (total COs; filled circles) and C (VDE-resistant COs; open circles). Data are from a single experiment. DOI: http://dx.doi.org/10.7554/eLife.19669.004
    Figure Legend Snippet: Spo11-initiated events at the two insert loci. ( A ) Spo11-catalyzed DSBs are more frequent at HIS4 that at URA3 . Left—Southern blots of Eco RI digests of DNA from vde∆ strains, probed with pBR322 sequences, showing Spo11-DSBs in the Parent 2 insert (see Figure 1 ) in resection/repair-deficient sae2∆ mutant strains. Right—location of DSBs and probe and DSB frequencies (average of 7 and 8 hr samples from a single experiment; error bars represent range). Spo11-DSBs in the Parent 1 inserts at HIS4 and URA3 were at different locations within the insert, but displayed similar ratios between the two loci (data not shown). ( B ) Southern blots of Hin dIII digests of DNA from vde∆ strains, to detect total Spo11-initiated crossovers. ( C ) Southern blots of Hin dIII-VDE double digests of the same samples, to determine the background contribution of Spo11-initiated COs in subsequent experiments measuring VDE-initiated COs, which will be VDE-resistant due to conversion of the VRS site to VRS103 . Probes were as shown in Figure 1 . ( D ) Quantification of data in panels B (total COs; filled circles) and C (VDE-resistant COs; open circles). Data are from a single experiment. DOI: http://dx.doi.org/10.7554/eLife.19669.004

    Techniques Used: Mutagenesis

    70–80% of VDE-DSBs are repaired. ( A ) Fraction of inserts remaining, calculated using Hin dIII digests (see Figure 1 ). For the arg4-VRS103 insert, the ratio (Parent 2 + CO2)/ (0.5 x LC) was calculated at 9 hr, and was then normalized to the 0 hr value. For the arg4-VRS insert, a similar calculation was made: (Parent 1 + NCO + CO1)/(0.5 x LC) ( B ) Relative recovery of interhomolog recombination products, calculated using Hin dIII-VDE double digests (see Figure 1 ). The sum of CO (average of CO1 and CO2) and NCO frequencies was divided by the frequency of total DSBs, as calculated in Figure 2A . Data are the average of two independent experiments; error bars represent range. DOI: http://dx.doi.org/10.7554/eLife.19669.006
    Figure Legend Snippet: 70–80% of VDE-DSBs are repaired. ( A ) Fraction of inserts remaining, calculated using Hin dIII digests (see Figure 1 ). For the arg4-VRS103 insert, the ratio (Parent 2 + CO2)/ (0.5 x LC) was calculated at 9 hr, and was then normalized to the 0 hr value. For the arg4-VRS insert, a similar calculation was made: (Parent 1 + NCO + CO1)/(0.5 x LC) ( B ) Relative recovery of interhomolog recombination products, calculated using Hin dIII-VDE double digests (see Figure 1 ). The sum of CO (average of CO1 and CO2) and NCO frequencies was divided by the frequency of total DSBs, as calculated in Figure 2A . Data are the average of two independent experiments; error bars represent range. DOI: http://dx.doi.org/10.7554/eLife.19669.006

    Techniques Used:

    Southern blots of Hin dIII and Hin dIII-VDE digests of DNA from HIS4 insert-containing strains (top) and from URA3 insert-contaning strains (bottom). Probes and gel labels are as in Figure 1 ; JM—joint molecule recombination intermediates. DOI: http://dx.doi.org/10.7554/eLife.19669.009
    Figure Legend Snippet: Southern blots of Hin dIII and Hin dIII-VDE digests of DNA from HIS4 insert-containing strains (top) and from URA3 insert-contaning strains (bottom). Probes and gel labels are as in Figure 1 ; JM—joint molecule recombination intermediates. DOI: http://dx.doi.org/10.7554/eLife.19669.009

    Techniques Used:

    Southern blots of Hin dIII and Hin dIII-VDE digests of DNA from HIS4 insert-containing strains (top) and from URA3 insert-contaning strains (bottom). Gel labels are as in Figure 1 ; JM—joint molecule recombination intermediates. In the gel with Hin DIII digests of samples from a pch2∆ mm4-mn yen1∆ slx1∆ strain with inserts at URA3 , the 9 hr sample was originally loaded between the 4 and 5 hr samples; this image was cut and spliced as indicated by vertical lines for presentation purposes. DOI: http://dx.doi.org/10.7554/eLife.19669.012
    Figure Legend Snippet: Southern blots of Hin dIII and Hin dIII-VDE digests of DNA from HIS4 insert-containing strains (top) and from URA3 insert-contaning strains (bottom). Gel labels are as in Figure 1 ; JM—joint molecule recombination intermediates. In the gel with Hin DIII digests of samples from a pch2∆ mm4-mn yen1∆ slx1∆ strain with inserts at URA3 , the 9 hr sample was originally loaded between the 4 and 5 hr samples; this image was cut and spliced as indicated by vertical lines for presentation purposes. DOI: http://dx.doi.org/10.7554/eLife.19669.012

    Techniques Used:

    2) Product Images from "Plant X-tender: An extension of the AssemblX system for the assembly and expression of multigene constructs in plants"

    Article Title: Plant X-tender: An extension of the AssemblX system for the assembly and expression of multigene constructs in plants

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0190526

    Design of Plant X-tender expression vectors. Vector pCAMBIA 1300 (A) or Gateway vectors (pK7WG, pH7WG or pB7WG) (B) were used as a backbone. (A) I- Sce I–A0– Hin dIII– ccd B– Hin dIII–B0–I- Sce I cassette was introduced into the MCS region of pCAMBIA1300 by overlap-based cloning methods after backbone digestion with Bam HI and Hin dIII to obtain pCAMBIA_ASX. (B) T35S–AttR2– ccd B–AttR1 cassette was released from the Gateway plasmid backbone by digestion with Xba I and Sac I and replaced with a I- Sce I–A0– Hin dIII– ccd B– Hin dIII–B0–I- Sce I cassette by overlap-based cloning methods to obtain pK7WG_ASX, pH7WG_ASX or pB7WG_ASX. MCS: multiple cloning site, A0/B0: homology regions, Kan: selection marker conferring kanamycin resistance in E . coli and A . tumefaciens , Spec: selection marker conferring spectinomycin resistance in E . coli and A . tumefaciens , Hyg: selection marker conferring hygromycin resistance in plants, R: selection marker conferring resistance in plants (kanamycin resistance in pK7WG, hygromycin resistance in pH7WG, herbicide glufosinate-ammonium resistance in pB7WG), LB: left border of T-DNA, RB: right border of T-DNA, ccd B: bacterial suicide gene, Hin dIII, I- Sce I, Bam HI, Xba I, Sac I: restriction enzyme recognition sites, AttR1/AttR2: Gateway cloning recombination sites, T35S: cauliflower mosaic virus CaMV 35S terminator, SLiCE: Seamless ligation cloning extract cloning method, HiFi: NEBuilder HiFi DNA assembly method, Gibson: Gibson DNA assembly method.
    Figure Legend Snippet: Design of Plant X-tender expression vectors. Vector pCAMBIA 1300 (A) or Gateway vectors (pK7WG, pH7WG or pB7WG) (B) were used as a backbone. (A) I- Sce I–A0– Hin dIII– ccd B– Hin dIII–B0–I- Sce I cassette was introduced into the MCS region of pCAMBIA1300 by overlap-based cloning methods after backbone digestion with Bam HI and Hin dIII to obtain pCAMBIA_ASX. (B) T35S–AttR2– ccd B–AttR1 cassette was released from the Gateway plasmid backbone by digestion with Xba I and Sac I and replaced with a I- Sce I–A0– Hin dIII– ccd B– Hin dIII–B0–I- Sce I cassette by overlap-based cloning methods to obtain pK7WG_ASX, pH7WG_ASX or pB7WG_ASX. MCS: multiple cloning site, A0/B0: homology regions, Kan: selection marker conferring kanamycin resistance in E . coli and A . tumefaciens , Spec: selection marker conferring spectinomycin resistance in E . coli and A . tumefaciens , Hyg: selection marker conferring hygromycin resistance in plants, R: selection marker conferring resistance in plants (kanamycin resistance in pK7WG, hygromycin resistance in pH7WG, herbicide glufosinate-ammonium resistance in pB7WG), LB: left border of T-DNA, RB: right border of T-DNA, ccd B: bacterial suicide gene, Hin dIII, I- Sce I, Bam HI, Xba I, Sac I: restriction enzyme recognition sites, AttR1/AttR2: Gateway cloning recombination sites, T35S: cauliflower mosaic virus CaMV 35S terminator, SLiCE: Seamless ligation cloning extract cloning method, HiFi: NEBuilder HiFi DNA assembly method, Gibson: Gibson DNA assembly method.

    Techniques Used: Expressing, Plasmid Preparation, Clone Assay, Selection, Marker, Ligation

    3) Product Images from "Modulation of RNase E Activity by Alternative RNA Binding Sites"

    Article Title: Modulation of RNase E Activity by Alternative RNA Binding Sites

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0090610

    Effects of Y25A and Q36R on the catalytic activity of RNase E in vivo and in vitro . (A) Plasmid copy number of pNRNE4, pNRNE4-Q36R and pNRNE4-Y25A in KSL2000. Plasmids were purified from KSL2000 cells harboring pNRNE4, pNRNE4-Q36R or pNRNE4-Y25A and were digested with Hin dIII, which has a unique cleavage site in all of the plasmids tested. Plasmid copy number was calculated relative to the concurrent presence of the pSC101 derivative (pBAD-RNE), which replicates independently of Rne, by measuring the molar ratio of the ColE1-type plasmid to the pBAD-RNE plasmid. (B) Growth characteristics of KSL2003 cells expressing wild-type N-Rne or the Q36R or Y25A mutant proteins. Growth of KSL2003 cells harboring pLAC-RNE2, pLAC-RNE2-Q36R, or pLAC-RNE2-Y25A was measured individually on LB-agar plates containing 1.0 to 1000 µM IPTG. Numbers on the top indicate the number of bacterial cells in each spot. (C) Plasmid copy number of pET28a in KSL2003. Plasmids were purified from KSL2003, KSL2003-Q36R or KSL2003-Y25A cells harboring pET28a and digested with Hin dIII, which has a unique cleavage site in all the plasmids tested. Plasmid copy number was calculated relative to the concurrent presence of the pSC101 derivative (pLAC-RNE2, pLAC-RNE2-Q36R or pLAC-RNE2-Y25A) by measuring the molar ratio of the ColE1-type plasmid to the pSC101-derived plasmid. (D) Expression profiles of Rne and mutant proteins in KSL2003. The membrane probed with an anti-Rne polyclonal antibody was stripped and reprobed with an anti-S1 polyclonal antibody to provide an internal standard. The relative abundance of protein was quantified by setting the amount of wild-type Rne to 1. KSL2003 cells were grown in LB medium containing 10 µM IPTG. (E) In vitro cleavage of p-BR13 by wild-type N-Rne, Q36R and Y25A mutant proteins. Two pmol of 5′ end-labeled p-BR13 was incubated with 1 pmol of purified wild-type N-Rne or Q36R or Y25A mutant protein in 20 µl of cleavage buffer at 37°C. Samples were removed at each indicated time point and mixed with an equal volume of loading buffer. Samples were denatured at 65°C for 5 min and loaded onto 15% polyacrylamide gel containing 8 M urea. The radioactivity in each band was quantified using a phosphorimager and OptiQuant software.
    Figure Legend Snippet: Effects of Y25A and Q36R on the catalytic activity of RNase E in vivo and in vitro . (A) Plasmid copy number of pNRNE4, pNRNE4-Q36R and pNRNE4-Y25A in KSL2000. Plasmids were purified from KSL2000 cells harboring pNRNE4, pNRNE4-Q36R or pNRNE4-Y25A and were digested with Hin dIII, which has a unique cleavage site in all of the plasmids tested. Plasmid copy number was calculated relative to the concurrent presence of the pSC101 derivative (pBAD-RNE), which replicates independently of Rne, by measuring the molar ratio of the ColE1-type plasmid to the pBAD-RNE plasmid. (B) Growth characteristics of KSL2003 cells expressing wild-type N-Rne or the Q36R or Y25A mutant proteins. Growth of KSL2003 cells harboring pLAC-RNE2, pLAC-RNE2-Q36R, or pLAC-RNE2-Y25A was measured individually on LB-agar plates containing 1.0 to 1000 µM IPTG. Numbers on the top indicate the number of bacterial cells in each spot. (C) Plasmid copy number of pET28a in KSL2003. Plasmids were purified from KSL2003, KSL2003-Q36R or KSL2003-Y25A cells harboring pET28a and digested with Hin dIII, which has a unique cleavage site in all the plasmids tested. Plasmid copy number was calculated relative to the concurrent presence of the pSC101 derivative (pLAC-RNE2, pLAC-RNE2-Q36R or pLAC-RNE2-Y25A) by measuring the molar ratio of the ColE1-type plasmid to the pSC101-derived plasmid. (D) Expression profiles of Rne and mutant proteins in KSL2003. The membrane probed with an anti-Rne polyclonal antibody was stripped and reprobed with an anti-S1 polyclonal antibody to provide an internal standard. The relative abundance of protein was quantified by setting the amount of wild-type Rne to 1. KSL2003 cells were grown in LB medium containing 10 µM IPTG. (E) In vitro cleavage of p-BR13 by wild-type N-Rne, Q36R and Y25A mutant proteins. Two pmol of 5′ end-labeled p-BR13 was incubated with 1 pmol of purified wild-type N-Rne or Q36R or Y25A mutant protein in 20 µl of cleavage buffer at 37°C. Samples were removed at each indicated time point and mixed with an equal volume of loading buffer. Samples were denatured at 65°C for 5 min and loaded onto 15% polyacrylamide gel containing 8 M urea. The radioactivity in each band was quantified using a phosphorimager and OptiQuant software.

    Techniques Used: Activity Assay, In Vivo, In Vitro, Plasmid Preparation, Purification, Expressing, Mutagenesis, Derivative Assay, Labeling, Incubation, Radioactivity, Software

    4) Product Images from "A nematode effector protein similar to annexins in host plants"

    Article Title: A nematode effector protein similar to annexins in host plants

    Journal: Journal of Experimental Botany

    doi: 10.1093/jxb/erp293

    Genomic DNA of Heterodera schachtii and H. glycines digested with Bam HI (B) and Hin dIII (H) was hybridized on blots with a 162 bp Hg4F01 DIG-labelled cDNA probe and revealed two potential 4F01 annexin-like family members in each cyst nematode species. M, molecular weight marker shown in Kb.
    Figure Legend Snippet: Genomic DNA of Heterodera schachtii and H. glycines digested with Bam HI (B) and Hin dIII (H) was hybridized on blots with a 162 bp Hg4F01 DIG-labelled cDNA probe and revealed two potential 4F01 annexin-like family members in each cyst nematode species. M, molecular weight marker shown in Kb.

    Techniques Used: Molecular Weight, Marker

    5) Product Images from "Construction and Characterization of a Bacterial Artificial Chromosome Library for the Hexaploid Wheat Line 92R137"

    Article Title: Construction and Characterization of a Bacterial Artificial Chromosome Library for the Hexaploid Wheat Line 92R137

    Journal: BioMed Research International

    doi: 10.1155/2014/845806

    Partial digestion of DNA in half plugs. Lanes 1–8 contain DNA samples digested with restriction enzyme at the increasingly higher concentrations. (a) Partial digestions of half DNA plugs with serial dilutions of Hin dIII at 37°C for 30 min. (b) Partial digestions of half DNA plugs with serial dilutions of Bam HI at 37°C for 30 min. Plug pieces were separated on 1% agarose gel in 0.5x TBE and run in the CHEF DR-III System (Bio-Rad) at 6 V/cm, 1–50 s switch time, linear ramp, 120° angle at 14°C for 18 h. M: the λ PFG marker.
    Figure Legend Snippet: Partial digestion of DNA in half plugs. Lanes 1–8 contain DNA samples digested with restriction enzyme at the increasingly higher concentrations. (a) Partial digestions of half DNA plugs with serial dilutions of Hin dIII at 37°C for 30 min. (b) Partial digestions of half DNA plugs with serial dilutions of Bam HI at 37°C for 30 min. Plug pieces were separated on 1% agarose gel in 0.5x TBE and run in the CHEF DR-III System (Bio-Rad) at 6 V/cm, 1–50 s switch time, linear ramp, 120° angle at 14°C for 18 h. M: the λ PFG marker.

    Techniques Used: Agarose Gel Electrophoresis, Marker

    6) Product Images from "An enhanced method for sequence walking and paralog mining: TOPO® Vector-Ligation PCR"

    Article Title: An enhanced method for sequence walking and paralog mining: TOPO® Vector-Ligation PCR

    Journal: BMC Research Notes

    doi: 10.1186/1756-0500-3-61

    Extension of sequence from clone 143-2-1 in the 5' (A) and 3' (B) directions . First round TVL-PCR products (top) were generated from templates prepared using Hin dIII (H), Eco RI (E), or Bam HI (B) digestion of genomic DNA. First round products were used as templates to generate second round products (bottom). Boxes (bottom gels) indicate the locations of the second round TVL-PCR products that proved to be the desired products.
    Figure Legend Snippet: Extension of sequence from clone 143-2-1 in the 5' (A) and 3' (B) directions . First round TVL-PCR products (top) were generated from templates prepared using Hin dIII (H), Eco RI (E), or Bam HI (B) digestion of genomic DNA. First round products were used as templates to generate second round products (bottom). Boxes (bottom gels) indicate the locations of the second round TVL-PCR products that proved to be the desired products.

    Techniques Used: Sequencing, Polymerase Chain Reaction, Generated

    7) Product Images from "An Agrobacterium tumefaciens Strain with Gamma-Aminobutyric Acid Transaminase Activity Shows an Enhanced Genetic Transformation Ability in Plants"

    Article Title: An Agrobacterium tumefaciens Strain with Gamma-Aminobutyric Acid Transaminase Activity Shows an Enhanced Genetic Transformation Ability in Plants

    Journal: Scientific Reports

    doi: 10.1038/srep42649

    GABA transaminase activity was introduced into A. tumefaciens . ( A ) Amino acid sequence comparison of GABA transaminase from E. coli K12 (gabTSCA772438), P. syringae pv. tomato DC3000 (gabT1: gabT1-PSPTO0259, gabT2: gabT2-PSPTO0301), P. aeruginosa PAO1 (gabT-PA0266). “ClustalW2” ( http://www.ebi.ac.uk/Tools/msa/clustalw2/ ) and “Jalview” ( http://www.jalview.org ) were used for calculation of amino acid multiple sequence alignment and display the alignment result, respectively. The residues were coloured according to their physicochemical properties as follows; Aliphatic/Hydrophobic, Aromatic, Positive, Negative, Hydrophilic, Conformationally Special and Cystein. Arrowheads indicate 3 of the 12 invariant amino acid residues among 16 aminotransferases with highly homologous peptides. The red box indicates a conserved motif (Ser [or Thr]-X-X-Lys) in the pyridoxalphosphate-binding peptide of aspartate aminotransferase (AAT) and histidinol-phosphate transaminases. The blue box indicates near identity between the homologous peptides of the histidinol-phosphate transaminases from E. coli K12 and Saccharomyces cerevisiae . ( B ) Construction of a plasmid for the expression of GABA transaminase ( gabT ) in A. tumefaciens. Hin dIII and Xba I fragments (ca. 1.6 kb) containing the GABA transaminase gene from E. coli K12 were ligated into the Hin dIII and Xba I sites of the broad-host-range plasmid pBBR1MCS-5, resulting in pBBR gabT . The expression of the GABA transaminase gene gabT was under the control of the lac promoter. MCS: multiple cloning site. ( C ) Detection of GABA activity in A. tumefaciens . Glutamic acid accumulation in the reaction buffer was measured according to the method of Akihiro et al . 48 . The open and closed circles indicate A. tumefaciens GV2260 (pBBR gabT , pIG121-Hm) and A. tumefaciens GV2260 (pBBR1MCS-5, pIG121-Hm), respectively. Bars represent the standard deviation (n = 3).
    Figure Legend Snippet: GABA transaminase activity was introduced into A. tumefaciens . ( A ) Amino acid sequence comparison of GABA transaminase from E. coli K12 (gabTSCA772438), P. syringae pv. tomato DC3000 (gabT1: gabT1-PSPTO0259, gabT2: gabT2-PSPTO0301), P. aeruginosa PAO1 (gabT-PA0266). “ClustalW2” ( http://www.ebi.ac.uk/Tools/msa/clustalw2/ ) and “Jalview” ( http://www.jalview.org ) were used for calculation of amino acid multiple sequence alignment and display the alignment result, respectively. The residues were coloured according to their physicochemical properties as follows; Aliphatic/Hydrophobic, Aromatic, Positive, Negative, Hydrophilic, Conformationally Special and Cystein. Arrowheads indicate 3 of the 12 invariant amino acid residues among 16 aminotransferases with highly homologous peptides. The red box indicates a conserved motif (Ser [or Thr]-X-X-Lys) in the pyridoxalphosphate-binding peptide of aspartate aminotransferase (AAT) and histidinol-phosphate transaminases. The blue box indicates near identity between the homologous peptides of the histidinol-phosphate transaminases from E. coli K12 and Saccharomyces cerevisiae . ( B ) Construction of a plasmid for the expression of GABA transaminase ( gabT ) in A. tumefaciens. Hin dIII and Xba I fragments (ca. 1.6 kb) containing the GABA transaminase gene from E. coli K12 were ligated into the Hin dIII and Xba I sites of the broad-host-range plasmid pBBR1MCS-5, resulting in pBBR gabT . The expression of the GABA transaminase gene gabT was under the control of the lac promoter. MCS: multiple cloning site. ( C ) Detection of GABA activity in A. tumefaciens . Glutamic acid accumulation in the reaction buffer was measured according to the method of Akihiro et al . 48 . The open and closed circles indicate A. tumefaciens GV2260 (pBBR gabT , pIG121-Hm) and A. tumefaciens GV2260 (pBBR1MCS-5, pIG121-Hm), respectively. Bars represent the standard deviation (n = 3).

    Techniques Used: Activity Assay, Sequencing, Binding Assay, Plasmid Preparation, Expressing, Clone Assay, Standard Deviation

    8) Product Images from "Plant X-tender: An extension of the AssemblX system for the assembly and expression of multigene constructs in plants"

    Article Title: Plant X-tender: An extension of the AssemblX system for the assembly and expression of multigene constructs in plants

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0190526

    Design of Plant X-tender expression vectors. Vector pCAMBIA 1300 (A) or Gateway vectors (pK7WG, pH7WG or pB7WG) (B) were used as a backbone. (A) I- Sce I–A0– Hin dIII– ccd B– Hin dIII–B0–I- Sce I cassette was introduced into the MCS region of pCAMBIA1300 by overlap-based cloning methods after backbone digestion with Bam HI and Hin dIII to obtain pCAMBIA_ASX. (B) T35S–AttR2– ccd B–AttR1 cassette was released from the Gateway plasmid backbone by digestion with Xba I and Sac I and replaced with a I- Sce I–A0– Hin dIII– ccd B– Hin dIII–B0–I- Sce I cassette by overlap-based cloning methods to obtain pK7WG_ASX, pH7WG_ASX or pB7WG_ASX. MCS: multiple cloning site, A0/B0: homology regions, Kan: selection marker conferring kanamycin resistance in E . coli and A . tumefaciens , Spec: selection marker conferring spectinomycin resistance in E . coli and A . tumefaciens , Hyg: selection marker conferring hygromycin resistance in plants, R: selection marker conferring resistance in plants (kanamycin resistance in pK7WG, hygromycin resistance in pH7WG, herbicide glufosinate-ammonium resistance in pB7WG), LB: left border of T-DNA, RB: right border of T-DNA, ccd B: bacterial suicide gene, Hin dIII, I- Sce I, Bam HI, Xba I, Sac I: restriction enzyme recognition sites, AttR1/AttR2: Gateway cloning recombination sites, T35S: cauliflower mosaic virus CaMV 35S terminator, SLiCE: Seamless ligation cloning extract cloning method, HiFi: NEBuilder HiFi DNA assembly method, Gibson: Gibson DNA assembly method.
    Figure Legend Snippet: Design of Plant X-tender expression vectors. Vector pCAMBIA 1300 (A) or Gateway vectors (pK7WG, pH7WG or pB7WG) (B) were used as a backbone. (A) I- Sce I–A0– Hin dIII– ccd B– Hin dIII–B0–I- Sce I cassette was introduced into the MCS region of pCAMBIA1300 by overlap-based cloning methods after backbone digestion with Bam HI and Hin dIII to obtain pCAMBIA_ASX. (B) T35S–AttR2– ccd B–AttR1 cassette was released from the Gateway plasmid backbone by digestion with Xba I and Sac I and replaced with a I- Sce I–A0– Hin dIII– ccd B– Hin dIII–B0–I- Sce I cassette by overlap-based cloning methods to obtain pK7WG_ASX, pH7WG_ASX or pB7WG_ASX. MCS: multiple cloning site, A0/B0: homology regions, Kan: selection marker conferring kanamycin resistance in E . coli and A . tumefaciens , Spec: selection marker conferring spectinomycin resistance in E . coli and A . tumefaciens , Hyg: selection marker conferring hygromycin resistance in plants, R: selection marker conferring resistance in plants (kanamycin resistance in pK7WG, hygromycin resistance in pH7WG, herbicide glufosinate-ammonium resistance in pB7WG), LB: left border of T-DNA, RB: right border of T-DNA, ccd B: bacterial suicide gene, Hin dIII, I- Sce I, Bam HI, Xba I, Sac I: restriction enzyme recognition sites, AttR1/AttR2: Gateway cloning recombination sites, T35S: cauliflower mosaic virus CaMV 35S terminator, SLiCE: Seamless ligation cloning extract cloning method, HiFi: NEBuilder HiFi DNA assembly method, Gibson: Gibson DNA assembly method.

    Techniques Used: Expressing, Plasmid Preparation, Clone Assay, Selection, Marker, Ligation

    9) Product Images from "The mismatch repair and meiotic recombination endonuclease Mlh1-Mlh3 is activated by polymer formation and can cleave DNA substrates in trans"

    Article Title: The mismatch repair and meiotic recombination endonuclease Mlh1-Mlh3 is activated by polymer formation and can cleave DNA substrates in trans

    Journal: PLoS Biology

    doi: 10.1371/journal.pbio.2001164

    Mlh1-Mlh3’s endonuclease activity requires a continuous substrate and increases as substrate size increases. (A) Denaturing agarose analysis of yeast Mlh1-Mlh3 nicking on circular pUC18 (linearized prior to gel loading) (2.7 kb; black), Hin dIII linearized pUC18 (red), and Hin dIII linearized pUC18 with streptavidin (SA) bound to ends (blue). Migration of linearized substrate (l) is indicated. (B) Average of two separate experiments: fraction nicked defined as fraction of substrate lost plotted against yeast Mlh1-Mlh3 concentration; error bars represent the standard deviation between three experiments. (C) Top: native agarose gel electrophoresis analysis of yeast Mlh1-Mlh3 (150 nM) endonuclease activity on circular substrate ranging from 2.7 kb to 12 kb. The concentration of nucleotide in each reaction is 15 μM. (D) Quantification of nicking in lanes 4, 7, 10, and 13 in C averaged from three separate experiments. Error bars indicate standard deviation. (E) Denaturing agarose analysis of yeast Mlh1-Mlh3 nicking on 12-kb circular DNA (black) and Hin dIII linearized 12 kb substrate (red). (F) Average of three separate experiments; error bars represent standard deviation. All nicking reactions were carried out for 60 min.
    Figure Legend Snippet: Mlh1-Mlh3’s endonuclease activity requires a continuous substrate and increases as substrate size increases. (A) Denaturing agarose analysis of yeast Mlh1-Mlh3 nicking on circular pUC18 (linearized prior to gel loading) (2.7 kb; black), Hin dIII linearized pUC18 (red), and Hin dIII linearized pUC18 with streptavidin (SA) bound to ends (blue). Migration of linearized substrate (l) is indicated. (B) Average of two separate experiments: fraction nicked defined as fraction of substrate lost plotted against yeast Mlh1-Mlh3 concentration; error bars represent the standard deviation between three experiments. (C) Top: native agarose gel electrophoresis analysis of yeast Mlh1-Mlh3 (150 nM) endonuclease activity on circular substrate ranging from 2.7 kb to 12 kb. The concentration of nucleotide in each reaction is 15 μM. (D) Quantification of nicking in lanes 4, 7, 10, and 13 in C averaged from three separate experiments. Error bars indicate standard deviation. (E) Denaturing agarose analysis of yeast Mlh1-Mlh3 nicking on 12-kb circular DNA (black) and Hin dIII linearized 12 kb substrate (red). (F) Average of three separate experiments; error bars represent standard deviation. All nicking reactions were carried out for 60 min.

    Techniques Used: Activity Assay, Migration, Concentration Assay, Standard Deviation, Agarose Gel Electrophoresis

    Mlh1-Mlh3 can make DSBs on large DNA substrates. (A) Mlh1-Mlh3 makes exclusively linear product from a closed circular substrate approximately 12 kb in size. Experiments were performed identical to the manner described in Fig 2C using the indicated sized plasmids. Red asterisk indicates location where nicked 12 kb plasmid migrates. (B) 15 μM total nucleotide 12 kb circular substrate was incubated with 300 nM Mlh1-Mlh3 for the indicated period of time. Plasmid linearized with Hin dIII was used as a marker for linear product in the lane 8 of the left panel. 12 kb plasmid treated with DNaseI is used as a marker for closed circular, linear, and nicked species in lane 1. Lane 9 is a negative control reaction in which Mlh1-Mlh3 was omitted to indicate migration of closed circular substrate. The plot indicates quantification of the representative gel shown. (C) Experiment is identical to that conducted in B, except 15 μM total nucleotide 2.7 kb circular substrate was used. The plot indicates quantification of the representative gel shown. (D) Native gel analysis of material in Fig 2E lanes 7–11. (E). DSBs made by Mlh1-Mlh3 can be religated. 12 kb linear product from Mlh1-Mlh3 endonuclease assay (“Mlh1-Mlh3 linear pdt”) was gel isolated and incubated with T4 polymerase where indicated with a + (lanes 12–13) followed by T4 DNA ligase where indicated with a + (lanes 11, 13). As controls, 12 kb closed circular plasmid was linearized with either Sca I or Hin dIII and religated (lanes 4–7) or linearized with Hin dIII and blunted with T4 polymerase followed by a religation step (lanes 8–9). Gel-isolated 12 kb closed circular DNA and Sca I-linearized DNA were ran in lanes 2–3 as migration markers.
    Figure Legend Snippet: Mlh1-Mlh3 can make DSBs on large DNA substrates. (A) Mlh1-Mlh3 makes exclusively linear product from a closed circular substrate approximately 12 kb in size. Experiments were performed identical to the manner described in Fig 2C using the indicated sized plasmids. Red asterisk indicates location where nicked 12 kb plasmid migrates. (B) 15 μM total nucleotide 12 kb circular substrate was incubated with 300 nM Mlh1-Mlh3 for the indicated period of time. Plasmid linearized with Hin dIII was used as a marker for linear product in the lane 8 of the left panel. 12 kb plasmid treated with DNaseI is used as a marker for closed circular, linear, and nicked species in lane 1. Lane 9 is a negative control reaction in which Mlh1-Mlh3 was omitted to indicate migration of closed circular substrate. The plot indicates quantification of the representative gel shown. (C) Experiment is identical to that conducted in B, except 15 μM total nucleotide 2.7 kb circular substrate was used. The plot indicates quantification of the representative gel shown. (D) Native gel analysis of material in Fig 2E lanes 7–11. (E). DSBs made by Mlh1-Mlh3 can be religated. 12 kb linear product from Mlh1-Mlh3 endonuclease assay (“Mlh1-Mlh3 linear pdt”) was gel isolated and incubated with T4 polymerase where indicated with a + (lanes 12–13) followed by T4 DNA ligase where indicated with a + (lanes 11, 13). As controls, 12 kb closed circular plasmid was linearized with either Sca I or Hin dIII and religated (lanes 4–7) or linearized with Hin dIII and blunted with T4 polymerase followed by a religation step (lanes 8–9). Gel-isolated 12 kb closed circular DNA and Sca I-linearized DNA were ran in lanes 2–3 as migration markers.

    Techniques Used: Plasmid Preparation, Incubation, Marker, Negative Control, Migration, Isolation

    An activated Mlh1-Mlh3-DNA complex can nick DNA in trans. 0.7 nM 7.2 kb closed circular M13mp18 phagemid and 1.8 nM 2.7 kb linear pUC18 substrate were incubated with 300 nM Mlh1-Mlh3 under standard endonuclease assay conditions either in isolation or within the same reaction as indicated. (A) Reaction products were run on an alkaline agarose gel. The 7.2 kb substrate was linearized with Hin dIII prior to loading in the alkaline agarose gel. The material in lane 4 and 5 was mixed and run as a control in lane 11 to demonstrate the readout for a negative result. The fraction of DNA nicked was measured by determining the band density in either the 7.2 kb linear band or the 2.7 kb linear band by subtracting the density in a region immediately above the band as background and comparing it to the band densities in the negative controls. (B) Prior to linearization with Hin dIII, 10 μL of each reaction was removed and run on a native agarose gel.
    Figure Legend Snippet: An activated Mlh1-Mlh3-DNA complex can nick DNA in trans. 0.7 nM 7.2 kb closed circular M13mp18 phagemid and 1.8 nM 2.7 kb linear pUC18 substrate were incubated with 300 nM Mlh1-Mlh3 under standard endonuclease assay conditions either in isolation or within the same reaction as indicated. (A) Reaction products were run on an alkaline agarose gel. The 7.2 kb substrate was linearized with Hin dIII prior to loading in the alkaline agarose gel. The material in lane 4 and 5 was mixed and run as a control in lane 11 to demonstrate the readout for a negative result. The fraction of DNA nicked was measured by determining the band density in either the 7.2 kb linear band or the 2.7 kb linear band by subtracting the density in a region immediately above the band as background and comparing it to the band densities in the negative controls. (B) Prior to linearization with Hin dIII, 10 μL of each reaction was removed and run on a native agarose gel.

    Techniques Used: Incubation, Isolation, Agarose Gel Electrophoresis

    10) Product Images from "Screening of a Fosmid Library of Marine Environmental Genomic DNA Fragments Reveals Four Clones Related to Members of the Order Planctomycetales"

    Article Title: Screening of a Fosmid Library of Marine Environmental Genomic DNA Fragments Reveals Four Clones Related to Members of the Order Planctomycetales

    Journal: Applied and Environmental Microbiology

    doi:

    Southern blot of Eco RI- and Hin dIII-digested fosmids hybridized with 32 P end-labeled Planctomycetales -specific 16S rRNA and universal rRNA oligonucleotides. Sizes (in base pairs) are indicated on the left. Lanes: 1, 5H12; 2, 6O13; 3, 6N14; 4, 7F15.
    Figure Legend Snippet: Southern blot of Eco RI- and Hin dIII-digested fosmids hybridized with 32 P end-labeled Planctomycetales -specific 16S rRNA and universal rRNA oligonucleotides. Sizes (in base pairs) are indicated on the left. Lanes: 1, 5H12; 2, 6O13; 3, 6N14; 4, 7F15.

    Techniques Used: Southern Blot, Labeling

    11) Product Images from "Regulation of the sol Locus Genes for Butanol and Acetone Formation in Clostridium acetobutylicum ATCC 824 by a Putative Transcriptional Repressor"

    Article Title: Regulation of the sol Locus Genes for Butanol and Acetone Formation in Clostridium acetobutylicum ATCC 824 by a Putative Transcriptional Repressor

    Journal: Journal of Bacteriology

    doi:

    Schematic representations (a) and results of PCR analysis (b) on wild-type (WT) C. acetobutylicum ATCC 824 and solR mutants B and H using primers (a) designed to amplify the junction between the vector portion of pO1X and the solR gene. For each gel in panel b, lane 1 contains Hin dIII-digested lambda marker, lane 2 contains the WT genomic template, lane 3 contains the solR mutant B template, and lane 4 contains the solR mutant H as the template. (Gel A) Extralong PCR using primers solR453 and solR1361 designed to amplify the complete insert. In lane 2, WT DNA shows an expected ∼0.9-kb band. This band is also seen, but much weaker, in both lanes 3 and 4. In addition, lane 4 contains a band with an apparent size of ∼7 kb (marked with an arrow), consistent with the presence of one insert of pO1X into solR of mutant H. (Gel B) PCR results using primers solR453 and Tc238. A band of ∼1.2 kb can be seen in lanes 3 and 4 with no product in lane 2. (Gel C) PCR results using primers Em373 and solR1361. A band of ∼2.1 kb is visible in lanes 3 and 4. Again, no product was observed with WT DNA (lane 2).
    Figure Legend Snippet: Schematic representations (a) and results of PCR analysis (b) on wild-type (WT) C. acetobutylicum ATCC 824 and solR mutants B and H using primers (a) designed to amplify the junction between the vector portion of pO1X and the solR gene. For each gel in panel b, lane 1 contains Hin dIII-digested lambda marker, lane 2 contains the WT genomic template, lane 3 contains the solR mutant B template, and lane 4 contains the solR mutant H as the template. (Gel A) Extralong PCR using primers solR453 and solR1361 designed to amplify the complete insert. In lane 2, WT DNA shows an expected ∼0.9-kb band. This band is also seen, but much weaker, in both lanes 3 and 4. In addition, lane 4 contains a band with an apparent size of ∼7 kb (marked with an arrow), consistent with the presence of one insert of pO1X into solR of mutant H. (Gel B) PCR results using primers solR453 and Tc238. A band of ∼1.2 kb can be seen in lanes 3 and 4 with no product in lane 2. (Gel C) PCR results using primers Em373 and solR1361. A band of ∼2.1 kb is visible in lanes 3 and 4. Again, no product was observed with WT DNA (lane 2).

    Techniques Used: Polymerase Chain Reaction, Plasmid Preparation, Marker, Mutagenesis

    12) Product Images from "Novel Method of Cell-Free In Vitro Synthesis of the Human Fibroblast Growth Factor 1 Gene"

    Article Title: Novel Method of Cell-Free In Vitro Synthesis of the Human Fibroblast Growth Factor 1 Gene

    Journal: Journal of Biomedicine and Biotechnology

    doi: 10.1155/2010/971340

    Confirmation of assembled plasmid containing the FGF1 gene . The pcDNA3.1/V5-His-TOPO-FGF1 plasmid was cleaved by Hin dIII and Xho I. Lane 1: DNA molecular weight markers. Lane 2: Two digestion fragments of 5433 bp and 561 bp.
    Figure Legend Snippet: Confirmation of assembled plasmid containing the FGF1 gene . The pcDNA3.1/V5-His-TOPO-FGF1 plasmid was cleaved by Hin dIII and Xho I. Lane 1: DNA molecular weight markers. Lane 2: Two digestion fragments of 5433 bp and 561 bp.

    Techniques Used: Plasmid Preparation, Molecular Weight

    13) Product Images from "Transduction of Cellular Sequence by a Human Immunodeficiency Virus Type 1-Derived Vector"

    Article Title: Transduction of Cellular Sequence by a Human Immunodeficiency Virus Type 1-Derived Vector

    Journal: Journal of Virology

    doi: 10.1128/JVI.75.23.11902-11906.2001

    Products and DNA sequence obtained from inverse PCR of the producer cell provirus. (A) Lanes: 1, DNA markers; 2, template-negative PCR control; 3, second-round inverse PCR products with Hin dIII-digested and self-ligated genomic DNA as templates. Band B1 was amplified from the HIV-1 self-ligated product. Band B2 was anticipated to contain the HIV-1 integration site and flanking producer cell genome sequence because of its strong intensity. Both bands were cloned and sequenced. (B) Flanking human genomic sequence at the 5′ end of the HIV-1 integration site. Bold underlined letters represent the first 48 bp of the HIV-1 provirus 5′ sequence. Italic letters represent the human chromosome 1 sequence adjacent to the integration site.
    Figure Legend Snippet: Products and DNA sequence obtained from inverse PCR of the producer cell provirus. (A) Lanes: 1, DNA markers; 2, template-negative PCR control; 3, second-round inverse PCR products with Hin dIII-digested and self-ligated genomic DNA as templates. Band B1 was amplified from the HIV-1 self-ligated product. Band B2 was anticipated to contain the HIV-1 integration site and flanking producer cell genome sequence because of its strong intensity. Both bands were cloned and sequenced. (B) Flanking human genomic sequence at the 5′ end of the HIV-1 integration site. Bold underlined letters represent the first 48 bp of the HIV-1 provirus 5′ sequence. Italic letters represent the human chromosome 1 sequence adjacent to the integration site.

    Techniques Used: Sequencing, Inverse PCR, Polymerase Chain Reaction, Amplification, Clone Assay

    14) Product Images from "Genetic Characteristics of Borrelia coriaceae Isolates from the Soft Tick Ornithodoros coriaceus (Acari: Argasidae)"

    Article Title: Genetic Characteristics of Borrelia coriaceae Isolates from the Soft Tick Ornithodoros coriaceus (Acari: Argasidae)

    Journal: Journal of Clinical Microbiology

    doi:

    Southern hybridization of Apa I (a)-, Bam HI (b)-, and Hin dIII (c)-digested DNAs of Borrelia isolates with digoxigenin-labeled Co53 DNA as a probe. Lanes: 1, CA434; 2, CA435; 3, Co53; 4, B. parkeri ; 5, CA4. Southern hybridization and detection of DNA which hybridized to the probe were done as recommended by Boehringer Mannheim for the digoxigenin DNA labeling and detection kit.
    Figure Legend Snippet: Southern hybridization of Apa I (a)-, Bam HI (b)-, and Hin dIII (c)-digested DNAs of Borrelia isolates with digoxigenin-labeled Co53 DNA as a probe. Lanes: 1, CA434; 2, CA435; 3, Co53; 4, B. parkeri ; 5, CA4. Southern hybridization and detection of DNA which hybridized to the probe were done as recommended by Boehringer Mannheim for the digoxigenin DNA labeling and detection kit.

    Techniques Used: Hybridization, Labeling, DNA Labeling

    15) Product Images from "Transgenic Cotton (Gossypium hirsutum L.) to Combat Weed Vagaries: Utility of an Apical Meristem-Targeted in planta Transformation Strategy to Introgress a Modified CP4-EPSPS Gene for Glyphosate Tolerance"

    Article Title: Transgenic Cotton (Gossypium hirsutum L.) to Combat Weed Vagaries: Utility of an Apical Meristem-Targeted in planta Transformation Strategy to Introgress a Modified CP4-EPSPS Gene for Glyphosate Tolerance

    Journal: Frontiers in Plant Science

    doi: 10.3389/fpls.2020.00768

    Molecular analysis of transformants in T1 generation. Polymerase chain reaction analysis of T1 generation plants for the amplification of panel (A) 500-bp right border region of T-DNA. (B) 750-bp nptII fragment. Lane M: 1 kb Ladder (Thermo Fisher Scientific); Lane B: Water Blank (all PCR components but without template DNA); Lane WT: wild type; Lanes 1–24 of panel (A) and 1–26 of panel (B): T1 generation transgenic plants; Lane P: positive control (modified CP4-EPSPS binary vector). (C) Western blot analysis of selected transgenic plants along with wild type. (i) Lanes 1–8: transgenic plants (1-1, 2-1, 3-4, 4-4, 22-5, 39-9, 42-1, 42-3). Lane L: Prestained protein ladder; Lane WT: wild type; (ii) Lane L: prestained protein ladder; Lane P: purified protein of CP4-EPSPS (30 ng); Lane WT: wild type; Lane 4: transgenic plant 4-5. (D) Genomic Southern analysis of T1 generation transgenic plants. (i) Hin dIII digested genomic DNA probed with DIG-labeled 750-bp nptII fragment. Lane L: Lambda DNA Hin dIII digest; Lane WT: wild-type DNA; Lanes 3–14: DNA of transgenic plants (46-1, 42-1, 42-3, 39-9, 22-5, 21-1, 17-2, 4-4, 4-5, 3-4, 2-1, 1-1); Lane P: positive control (10 pg DNA of modified CP4-EPSPS binary vector). (ii) Bam HI digested genomic DNA probed with 420-bp CP4-EPSPS gene specific fragment. Lane L: Lambda DNA Hin dIII digest; Lane 2: 4-4; Lane 3: 4-5; Lane 4: positive control (10-pg modified CP4-EPSPS vector DNA); Lane 5: wild-type DNA.
    Figure Legend Snippet: Molecular analysis of transformants in T1 generation. Polymerase chain reaction analysis of T1 generation plants for the amplification of panel (A) 500-bp right border region of T-DNA. (B) 750-bp nptII fragment. Lane M: 1 kb Ladder (Thermo Fisher Scientific); Lane B: Water Blank (all PCR components but without template DNA); Lane WT: wild type; Lanes 1–24 of panel (A) and 1–26 of panel (B): T1 generation transgenic plants; Lane P: positive control (modified CP4-EPSPS binary vector). (C) Western blot analysis of selected transgenic plants along with wild type. (i) Lanes 1–8: transgenic plants (1-1, 2-1, 3-4, 4-4, 22-5, 39-9, 42-1, 42-3). Lane L: Prestained protein ladder; Lane WT: wild type; (ii) Lane L: prestained protein ladder; Lane P: purified protein of CP4-EPSPS (30 ng); Lane WT: wild type; Lane 4: transgenic plant 4-5. (D) Genomic Southern analysis of T1 generation transgenic plants. (i) Hin dIII digested genomic DNA probed with DIG-labeled 750-bp nptII fragment. Lane L: Lambda DNA Hin dIII digest; Lane WT: wild-type DNA; Lanes 3–14: DNA of transgenic plants (46-1, 42-1, 42-3, 39-9, 22-5, 21-1, 17-2, 4-4, 4-5, 3-4, 2-1, 1-1); Lane P: positive control (10 pg DNA of modified CP4-EPSPS binary vector). (ii) Bam HI digested genomic DNA probed with 420-bp CP4-EPSPS gene specific fragment. Lane L: Lambda DNA Hin dIII digest; Lane 2: 4-4; Lane 3: 4-5; Lane 4: positive control (10-pg modified CP4-EPSPS vector DNA); Lane 5: wild-type DNA.

    Techniques Used: Polymerase Chain Reaction, Amplification, Transgenic Assay, Positive Control, Modification, Plasmid Preparation, Western Blot, Purification, Labeling, Lambda DNA Preparation

    16) Product Images from "Plant X-tender: An extension of the AssemblX system for the assembly and expression of multigene constructs in plants"

    Article Title: Plant X-tender: An extension of the AssemblX system for the assembly and expression of multigene constructs in plants

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0190526

    Functional evaluation of constructed vectors by cloning expression cassette p35S::H2BRFP_tNOS into Plant X-tender expression vectors. (A-F) Scheme of the cloning procedure. (A) Amplification of expression cassette from template plasmid using primers with appropriate 5’ and 3’ extensions to add A0 and AR homology regions. (B) Expression cassette assembly in Hin dIII restricted pL0A_0-R Level 0 vector by NEBuilder HiFi assembly method. (C) Release of expression cassette with flanking homology regions A0 and AR from Level 0 vector by Pme I digestion. (D) Assembly of expression cassette with flanking homology regions A0 and AR into Pac I digested pL1A-hc / pL1A-lc (A0/AR) Level 1 vector by TAR or NEBuilder HiFi. (E) Release of expression cassette flanked by URA3 yeast selection marker and homology regions A0 and B0 from Level 1 vector by I- Sce I digestion. (F) Assembly of expression cassette flanked by URA3 yeast selection marker and homology regions A0 and B0 into Plant X-tender expression vectors by SLiCE or NEBuilder HiFi. (G-I) Images of agroinfiltrated N . benthamiana leaves obtained by laser scanning confocal microscopy. Leaves were agroinfiltrated with agrobacteria containing pCAMBIA_ASX_cassette, pK7WG_ASX_cassette, pH7WG_ASX_cassette, pB7WG_ASX_cassette or empty agrobacteria (top to bottom). (G) Nuclear localisation of RFP. Fluorescence is represented as maximum projections of z-stacks. (H) Bright field. (I) Overlay of G with H. Scale bars are 100 μm. p35S: cauliflower mosaic virus CaMV 35S promoter, H2BRFP: histon sequence fused to red fluorescence protein (mRFP1), tNOS: nopaline synthase terminator, A0, AR, B0: homology regions, Rp: selection marker conferring resistance in plants (hygromycin in the case of pCAMBIA_ASX and pH7WG_ASX, kanamycin in the case of pK7WG_ASX, glufosinate-ammonium in the case of pB7WG_ASX), Re: selection marker conferring resistance in E . coli and A . tumefaciens (kanamycin in the case of pCAMBIA_ASX, spectinomycinin in the case of pK7WG_ASX, pH7WG_ASX and pB7WG_ASX), Amp: selection marker conferring ampicillin resistance in E . coli and A . tumefaciens , Kan: selection marker conferring kanamycin resistance in E . coli and A . tumefaciens , LB: left border of T-DNA, RB: right border of T-DNA, Hin dIII, I- Sce I, Pac I, Pme I: restriction enzyme recognition sites, URA3 : yeast selection marker, ccd B: bacterial suicide gene, SLiCE: Seamless ligation cloning extract cloning method, HiFi: NEBuilder HiFi DNA assembly method, Gibson: Gibson DNA assembly method, TAR: cloning based on transformation-associated recombination, PCR: Polymerase chain reaction, ASX: Plant X-tender expression vector.
    Figure Legend Snippet: Functional evaluation of constructed vectors by cloning expression cassette p35S::H2BRFP_tNOS into Plant X-tender expression vectors. (A-F) Scheme of the cloning procedure. (A) Amplification of expression cassette from template plasmid using primers with appropriate 5’ and 3’ extensions to add A0 and AR homology regions. (B) Expression cassette assembly in Hin dIII restricted pL0A_0-R Level 0 vector by NEBuilder HiFi assembly method. (C) Release of expression cassette with flanking homology regions A0 and AR from Level 0 vector by Pme I digestion. (D) Assembly of expression cassette with flanking homology regions A0 and AR into Pac I digested pL1A-hc / pL1A-lc (A0/AR) Level 1 vector by TAR or NEBuilder HiFi. (E) Release of expression cassette flanked by URA3 yeast selection marker and homology regions A0 and B0 from Level 1 vector by I- Sce I digestion. (F) Assembly of expression cassette flanked by URA3 yeast selection marker and homology regions A0 and B0 into Plant X-tender expression vectors by SLiCE or NEBuilder HiFi. (G-I) Images of agroinfiltrated N . benthamiana leaves obtained by laser scanning confocal microscopy. Leaves were agroinfiltrated with agrobacteria containing pCAMBIA_ASX_cassette, pK7WG_ASX_cassette, pH7WG_ASX_cassette, pB7WG_ASX_cassette or empty agrobacteria (top to bottom). (G) Nuclear localisation of RFP. Fluorescence is represented as maximum projections of z-stacks. (H) Bright field. (I) Overlay of G with H. Scale bars are 100 μm. p35S: cauliflower mosaic virus CaMV 35S promoter, H2BRFP: histon sequence fused to red fluorescence protein (mRFP1), tNOS: nopaline synthase terminator, A0, AR, B0: homology regions, Rp: selection marker conferring resistance in plants (hygromycin in the case of pCAMBIA_ASX and pH7WG_ASX, kanamycin in the case of pK7WG_ASX, glufosinate-ammonium in the case of pB7WG_ASX), Re: selection marker conferring resistance in E . coli and A . tumefaciens (kanamycin in the case of pCAMBIA_ASX, spectinomycinin in the case of pK7WG_ASX, pH7WG_ASX and pB7WG_ASX), Amp: selection marker conferring ampicillin resistance in E . coli and A . tumefaciens , Kan: selection marker conferring kanamycin resistance in E . coli and A . tumefaciens , LB: left border of T-DNA, RB: right border of T-DNA, Hin dIII, I- Sce I, Pac I, Pme I: restriction enzyme recognition sites, URA3 : yeast selection marker, ccd B: bacterial suicide gene, SLiCE: Seamless ligation cloning extract cloning method, HiFi: NEBuilder HiFi DNA assembly method, Gibson: Gibson DNA assembly method, TAR: cloning based on transformation-associated recombination, PCR: Polymerase chain reaction, ASX: Plant X-tender expression vector.

    Techniques Used: Functional Assay, Construct, Clone Assay, Expressing, Amplification, Plasmid Preparation, Selection, Marker, Confocal Microscopy, Fluorescence, Sequencing, Ligation, Transformation Assay, Polymerase Chain Reaction

    17) Product Images from "Atypical 16S rRNA Gene Copies in Ochrobactrum intermedium Strains Reveal a Large Genomic Rearrangement by Recombination between rrn Copies"

    Article Title: Atypical 16S rRNA Gene Copies in Ochrobactrum intermedium Strains Reveal a Large Genomic Rearrangement by Recombination between rrn Copies

    Journal: Journal of Bacteriology

    doi: 10.1128/JB.185.9.2901-2909.2003

    PFGE migration of undigested and I- Ceu I-digested genomic DNAs of O. intermedium strains ADV1 and ADV3. (A) Migration of high-molecular-weight fragments. Lanes 1 and 2, undigested DNA from strain ADV1 (lane 1) and strain ADV3 (lane 2); lanes 3 and 4, I- Ceu I-digested DNA from strain ADV1 (lane 3) and strain ADV3 (lane 4); Lanes Sc and Sp , Saccharomyces cerevisiae ( Sc ) and Schizosaccharomyces pombe ( Sp ) chromosomes (Bio-Rad) as molecular size markers. (B) Migration of low-molecular-weight I- Ceu I fragments. Lane 1, strain ADV1; lane 2, strain ADV3. Chromosomes and I- Ceu I digestion patterns of O. intermedium strains ADV2 and ADV4 to -7 are identical to patterns for strains ADV1 and ADV3, respectively (data not shown). A mixture of λ digested by Hin dIII, the λ concatemer, and Saccharomyces cerevisiae chromosomes was used as the molecular size marker (lane λ/ Sc ); the bands useful for the measure were, from the bottom, 27, 50, 100, 150, 200, 225, 250, and 285 kb.
    Figure Legend Snippet: PFGE migration of undigested and I- Ceu I-digested genomic DNAs of O. intermedium strains ADV1 and ADV3. (A) Migration of high-molecular-weight fragments. Lanes 1 and 2, undigested DNA from strain ADV1 (lane 1) and strain ADV3 (lane 2); lanes 3 and 4, I- Ceu I-digested DNA from strain ADV1 (lane 3) and strain ADV3 (lane 4); Lanes Sc and Sp , Saccharomyces cerevisiae ( Sc ) and Schizosaccharomyces pombe ( Sp ) chromosomes (Bio-Rad) as molecular size markers. (B) Migration of low-molecular-weight I- Ceu I fragments. Lane 1, strain ADV1; lane 2, strain ADV3. Chromosomes and I- Ceu I digestion patterns of O. intermedium strains ADV2 and ADV4 to -7 are identical to patterns for strains ADV1 and ADV3, respectively (data not shown). A mixture of λ digested by Hin dIII, the λ concatemer, and Saccharomyces cerevisiae chromosomes was used as the molecular size marker (lane λ/ Sc ); the bands useful for the measure were, from the bottom, 27, 50, 100, 150, 200, 225, 250, and 285 kb.

    Techniques Used: Migration, Molecular Weight, Marker

    Copy numbers of 16S rDNA (A) and 46-bp insertion (B) in the genome of O. intermedium. Shown are Southern blots of Hin dIII-digested genomic DNA from strains PR17/sat (lane 1), ADV1 (lane 2), ADV3 (lane 3), and ADV9 (lane 4) and reference strain LMG 3301 T (lane 5) hybridized with the 16S rDNA probe (A) and the 46-bp insertion probe (B). Strain ADV2 showed a hybridization profile identical to that of strain ADV1; strains ADV4 to -7 showed hybridization profiles identical to that of strain ADV3; strains ADV10, ADV11, ADV14, and ADV24 showed hybridization profiles identical to the strain LMG 3301 T profile (data not shown). Sizes of hybridizing fragments were calculated by using λ digested by Hin dIII as a molecular marker.
    Figure Legend Snippet: Copy numbers of 16S rDNA (A) and 46-bp insertion (B) in the genome of O. intermedium. Shown are Southern blots of Hin dIII-digested genomic DNA from strains PR17/sat (lane 1), ADV1 (lane 2), ADV3 (lane 3), and ADV9 (lane 4) and reference strain LMG 3301 T (lane 5) hybridized with the 16S rDNA probe (A) and the 46-bp insertion probe (B). Strain ADV2 showed a hybridization profile identical to that of strain ADV1; strains ADV4 to -7 showed hybridization profiles identical to that of strain ADV3; strains ADV10, ADV11, ADV14, and ADV24 showed hybridization profiles identical to the strain LMG 3301 T profile (data not shown). Sizes of hybridizing fragments were calculated by using λ digested by Hin dIII as a molecular marker.

    Techniques Used: Hybridization, Marker

    18) Product Images from "Genomics of Three New Bacteriophages Useful in the Biocontrol of Salmonella"

    Article Title: Genomics of Three New Bacteriophages Useful in the Biocontrol of Salmonella

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2016.00545

    Determination of genome ends of UAB_Phi20 phage after digestion with EcoRI enzyme . Genome of bacteriophage P22 digested with Eco RI was used as control. Arrows indicate the 4007-bp fragment containing the pac sequence. Lambda DNA digested with Hin dIII (M1) or Bst EII (M2) were used as molecular markers. Sizes (bp) are indicated on both sides of the image.
    Figure Legend Snippet: Determination of genome ends of UAB_Phi20 phage after digestion with EcoRI enzyme . Genome of bacteriophage P22 digested with Eco RI was used as control. Arrows indicate the 4007-bp fragment containing the pac sequence. Lambda DNA digested with Hin dIII (M1) or Bst EII (M2) were used as molecular markers. Sizes (bp) are indicated on both sides of the image.

    Techniques Used: Sequencing, Lambda DNA Preparation

    Time-limited digestion with Bal 31 exonuclease of UAB_Phi78 and UAB_Phi87 DNA followed by digestion with Hin dIII and Spe I, respectively . Arrows indicate the sequentially degraded DNA bands of 2200 and 2080 bp for UAB_Phi78 (A) and of 4322 and 2819 bp for UAB_Phi87 (B) . M: marker lanes containing a mixture of λ DNA digested with Bst EII and φX174 digested with Hin fI (M1), λ-DNA-digested Hin dIII (M2), and λ-DNA-digested Bst EII (M3). Sizes (bp) are indicated on the left side of the images.
    Figure Legend Snippet: Time-limited digestion with Bal 31 exonuclease of UAB_Phi78 and UAB_Phi87 DNA followed by digestion with Hin dIII and Spe I, respectively . Arrows indicate the sequentially degraded DNA bands of 2200 and 2080 bp for UAB_Phi78 (A) and of 4322 and 2819 bp for UAB_Phi87 (B) . M: marker lanes containing a mixture of λ DNA digested with Bst EII and φX174 digested with Hin fI (M1), λ-DNA-digested Hin dIII (M2), and λ-DNA-digested Bst EII (M3). Sizes (bp) are indicated on the left side of the images.

    Techniques Used: Marker

    19) Product Images from "Plant X-tender: An extension of the AssemblX system for the assembly and expression of multigene constructs in plants"

    Article Title: Plant X-tender: An extension of the AssemblX system for the assembly and expression of multigene constructs in plants

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0190526

    Functional evaluation of constructed vectors by cloning expression cassette p35S::H2BRFP_tNOS into Plant X-tender expression vectors. (A-F) Scheme of the cloning procedure. (A) Amplification of expression cassette from template plasmid using primers with appropriate 5’ and 3’ extensions to add A0 and AR homology regions. (B) Expression cassette assembly in Hin dIII restricted pL0A_0-R Level 0 vector by NEBuilder HiFi assembly method. (C) Release of expression cassette with flanking homology regions A0 and AR from Level 0 vector by Pme I digestion. (D) Assembly of expression cassette with flanking homology regions A0 and AR into Pac I digested pL1A-hc / pL1A-lc (A0/AR) Level 1 vector by TAR or NEBuilder HiFi. (E) Release of expression cassette flanked by URA3 yeast selection marker and homology regions A0 and B0 from Level 1 vector by I- Sce I digestion. (F) Assembly of expression cassette flanked by URA3 yeast selection marker and homology regions A0 and B0 into Plant X-tender expression vectors by SLiCE or NEBuilder HiFi. (G-I) Images of agroinfiltrated N . benthamiana leaves obtained by laser scanning confocal microscopy. Leaves were agroinfiltrated with agrobacteria containing pCAMBIA_ASX_cassette, pK7WG_ASX_cassette, pH7WG_ASX_cassette, pB7WG_ASX_cassette or empty agrobacteria (top to bottom). (G) Nuclear localisation of RFP. Fluorescence is represented as maximum projections of z-stacks. (H) Bright field. (I) Overlay of G with H. Scale bars are 100 μm. p35S: cauliflower mosaic virus CaMV 35S promoter, H2BRFP: histon sequence fused to red fluorescence protein (mRFP1), tNOS: nopaline synthase terminator, A0, AR, B0: homology regions, Rp: selection marker conferring resistance in plants (hygromycin in the case of pCAMBIA_ASX and pH7WG_ASX, kanamycin in the case of pK7WG_ASX, glufosinate-ammonium in the case of pB7WG_ASX), Re: selection marker conferring resistance in E . coli and A . tumefaciens (kanamycin in the case of pCAMBIA_ASX, spectinomycinin in the case of pK7WG_ASX, pH7WG_ASX and pB7WG_ASX), Amp: selection marker conferring ampicillin resistance in E . coli and A . tumefaciens , Kan: selection marker conferring kanamycin resistance in E . coli and A . tumefaciens , LB: left border of T-DNA, RB: right border of T-DNA, Hin dIII, I- Sce I, Pac I, Pme I: restriction enzyme recognition sites, URA3 : yeast selection marker, ccd B: bacterial suicide gene, SLiCE: Seamless ligation cloning extract cloning method, HiFi: NEBuilder HiFi DNA assembly method, Gibson: Gibson DNA assembly method, TAR: cloning based on transformation-associated recombination, PCR: Polymerase chain reaction, ASX: Plant X-tender expression vector.
    Figure Legend Snippet: Functional evaluation of constructed vectors by cloning expression cassette p35S::H2BRFP_tNOS into Plant X-tender expression vectors. (A-F) Scheme of the cloning procedure. (A) Amplification of expression cassette from template plasmid using primers with appropriate 5’ and 3’ extensions to add A0 and AR homology regions. (B) Expression cassette assembly in Hin dIII restricted pL0A_0-R Level 0 vector by NEBuilder HiFi assembly method. (C) Release of expression cassette with flanking homology regions A0 and AR from Level 0 vector by Pme I digestion. (D) Assembly of expression cassette with flanking homology regions A0 and AR into Pac I digested pL1A-hc / pL1A-lc (A0/AR) Level 1 vector by TAR or NEBuilder HiFi. (E) Release of expression cassette flanked by URA3 yeast selection marker and homology regions A0 and B0 from Level 1 vector by I- Sce I digestion. (F) Assembly of expression cassette flanked by URA3 yeast selection marker and homology regions A0 and B0 into Plant X-tender expression vectors by SLiCE or NEBuilder HiFi. (G-I) Images of agroinfiltrated N . benthamiana leaves obtained by laser scanning confocal microscopy. Leaves were agroinfiltrated with agrobacteria containing pCAMBIA_ASX_cassette, pK7WG_ASX_cassette, pH7WG_ASX_cassette, pB7WG_ASX_cassette or empty agrobacteria (top to bottom). (G) Nuclear localisation of RFP. Fluorescence is represented as maximum projections of z-stacks. (H) Bright field. (I) Overlay of G with H. Scale bars are 100 μm. p35S: cauliflower mosaic virus CaMV 35S promoter, H2BRFP: histon sequence fused to red fluorescence protein (mRFP1), tNOS: nopaline synthase terminator, A0, AR, B0: homology regions, Rp: selection marker conferring resistance in plants (hygromycin in the case of pCAMBIA_ASX and pH7WG_ASX, kanamycin in the case of pK7WG_ASX, glufosinate-ammonium in the case of pB7WG_ASX), Re: selection marker conferring resistance in E . coli and A . tumefaciens (kanamycin in the case of pCAMBIA_ASX, spectinomycinin in the case of pK7WG_ASX, pH7WG_ASX and pB7WG_ASX), Amp: selection marker conferring ampicillin resistance in E . coli and A . tumefaciens , Kan: selection marker conferring kanamycin resistance in E . coli and A . tumefaciens , LB: left border of T-DNA, RB: right border of T-DNA, Hin dIII, I- Sce I, Pac I, Pme I: restriction enzyme recognition sites, URA3 : yeast selection marker, ccd B: bacterial suicide gene, SLiCE: Seamless ligation cloning extract cloning method, HiFi: NEBuilder HiFi DNA assembly method, Gibson: Gibson DNA assembly method, TAR: cloning based on transformation-associated recombination, PCR: Polymerase chain reaction, ASX: Plant X-tender expression vector.

    Techniques Used: Functional Assay, Construct, Clone Assay, Expressing, Amplification, Plasmid Preparation, Selection, Marker, Confocal Microscopy, Fluorescence, Sequencing, Ligation, Transformation Assay, Polymerase Chain Reaction

    20) Product Images from "Conformation, Length, and Speed Measurements of Electrodynamically Stretched DNA in Nanochannels"

    Article Title: Conformation, Length, and Speed Measurements of Electrodynamically Stretched DNA in Nanochannels

    Journal: Biophysical Journal

    doi: 10.1529/biophysj.107.121020

    Photon count, length, speed, and size. Analysis of a sample containing a mixture of λ -bacteriophage DNA and its Hin dIII digest. In 2 min, 16,315 molecules were detected with an average speed ( A ) Distribution of the real length The peaks are shown fitted to nine modified Gaussians ( red ). ( B ) Real length versus number of basepairs. ( C ) Photon count (cnts) per molecule versus the number of basepairs. A linear fit yields a slope of ( D ) Distribution of photon counts cnts per molecule versus real length ( E ) Distribution of the photon counts per molecule. The peaks are fitted to eight modified Gaussians ( red ). The peaks in A and E are interpreted as the following DNA strand sizes, in number of basepairs: 125, 564, (2027 + 2322)/2, 4361, 6557, 9416, 23,130, 23,130 + 4361, 48,502. Whereas peaks 7 and 8 are resolved separately in A , they are combined in E . The values plotted in B and C are the mean values resulting from the Gaussian fits from A and E , respectively. ( F ) Distribution of molecule speed versus real length DNA speed is essentially constant with length, showing only a slight decrease for the shortest fragment.
    Figure Legend Snippet: Photon count, length, speed, and size. Analysis of a sample containing a mixture of λ -bacteriophage DNA and its Hin dIII digest. In 2 min, 16,315 molecules were detected with an average speed ( A ) Distribution of the real length The peaks are shown fitted to nine modified Gaussians ( red ). ( B ) Real length versus number of basepairs. ( C ) Photon count (cnts) per molecule versus the number of basepairs. A linear fit yields a slope of ( D ) Distribution of photon counts cnts per molecule versus real length ( E ) Distribution of the photon counts per molecule. The peaks are fitted to eight modified Gaussians ( red ). The peaks in A and E are interpreted as the following DNA strand sizes, in number of basepairs: 125, 564, (2027 + 2322)/2, 4361, 6557, 9416, 23,130, 23,130 + 4361, 48,502. Whereas peaks 7 and 8 are resolved separately in A , they are combined in E . The values plotted in B and C are the mean values resulting from the Gaussian fits from A and E , respectively. ( F ) Distribution of molecule speed versus real length DNA speed is essentially constant with length, showing only a slight decrease for the shortest fragment.

    Techniques Used: Modification

    21) Product Images from "Atypical 16S rRNA Gene Copies in Ochrobactrum intermedium Strains Reveal a Large Genomic Rearrangement by Recombination between rrn Copies"

    Article Title: Atypical 16S rRNA Gene Copies in Ochrobactrum intermedium Strains Reveal a Large Genomic Rearrangement by Recombination between rrn Copies

    Journal: Journal of Bacteriology

    doi: 10.1128/JB.185.9.2901-2909.2003

    PFGE migration of undigested and I- Ceu I-digested genomic DNAs of O. intermedium strains ADV1 and ADV3. (A) Migration of high-molecular-weight fragments. Lanes 1 and 2, undigested DNA from strain ADV1 (lane 1) and strain ADV3 (lane 2); lanes 3 and 4, I- Ceu I-digested DNA from strain ADV1 (lane 3) and strain ADV3 (lane 4); Lanes Sc and Sp , Saccharomyces cerevisiae ( Sc ) and Schizosaccharomyces pombe ( Sp ) chromosomes (Bio-Rad) as molecular size markers. (B) Migration of low-molecular-weight I- Ceu I fragments. Lane 1, strain ADV1; lane 2, strain ADV3. Chromosomes and I- Ceu I digestion patterns of O. intermedium strains ADV2 and ADV4 to -7 are identical to patterns for strains ADV1 and ADV3, respectively (data not shown). A mixture of λ digested by Hin dIII, the λ concatemer, and Saccharomyces cerevisiae chromosomes was used as the molecular size marker (lane λ/ Sc ); the bands useful for the measure were, from the bottom, 27, 50, 100, 150, 200, 225, 250, and 285 kb.
    Figure Legend Snippet: PFGE migration of undigested and I- Ceu I-digested genomic DNAs of O. intermedium strains ADV1 and ADV3. (A) Migration of high-molecular-weight fragments. Lanes 1 and 2, undigested DNA from strain ADV1 (lane 1) and strain ADV3 (lane 2); lanes 3 and 4, I- Ceu I-digested DNA from strain ADV1 (lane 3) and strain ADV3 (lane 4); Lanes Sc and Sp , Saccharomyces cerevisiae ( Sc ) and Schizosaccharomyces pombe ( Sp ) chromosomes (Bio-Rad) as molecular size markers. (B) Migration of low-molecular-weight I- Ceu I fragments. Lane 1, strain ADV1; lane 2, strain ADV3. Chromosomes and I- Ceu I digestion patterns of O. intermedium strains ADV2 and ADV4 to -7 are identical to patterns for strains ADV1 and ADV3, respectively (data not shown). A mixture of λ digested by Hin dIII, the λ concatemer, and Saccharomyces cerevisiae chromosomes was used as the molecular size marker (lane λ/ Sc ); the bands useful for the measure were, from the bottom, 27, 50, 100, 150, 200, 225, 250, and 285 kb.

    Techniques Used: Migration, Molecular Weight, Marker

    22) Product Images from "Construction and Characterization of a Bacterial Artificial Chromosome Library for the Hexaploid Wheat Line 92R137"

    Article Title: Construction and Characterization of a Bacterial Artificial Chromosome Library for the Hexaploid Wheat Line 92R137

    Journal: BioMed Research International

    doi: 10.1155/2014/845806

    Insert size distribution of randomly selected BAC clones including 453 clones constructed with Hin dIII (open bars) and 573 clones constructed with Bam HI (solid bars).
    Figure Legend Snippet: Insert size distribution of randomly selected BAC clones including 453 clones constructed with Hin dIII (open bars) and 573 clones constructed with Bam HI (solid bars).

    Techniques Used: BAC Assay, Clone Assay, Construct

    Partial digestion of DNA in half plugs. Lanes 1–8 contain DNA samples digested with restriction enzyme at the increasingly higher concentrations. (a) Partial digestions of half DNA plugs with serial dilutions of Hin dIII at 37°C for 30 min. (b) Partial digestions of half DNA plugs with serial dilutions of Bam HI at 37°C for 30 min. Plug pieces were separated on 1% agarose gel in 0.5x TBE and run in the CHEF DR-III System (Bio-Rad) at 6 V/cm, 1–50 s switch time, linear ramp, 120° angle at 14°C for 18 h. M: the λ PFG marker.
    Figure Legend Snippet: Partial digestion of DNA in half plugs. Lanes 1–8 contain DNA samples digested with restriction enzyme at the increasingly higher concentrations. (a) Partial digestions of half DNA plugs with serial dilutions of Hin dIII at 37°C for 30 min. (b) Partial digestions of half DNA plugs with serial dilutions of Bam HI at 37°C for 30 min. Plug pieces were separated on 1% agarose gel in 0.5x TBE and run in the CHEF DR-III System (Bio-Rad) at 6 V/cm, 1–50 s switch time, linear ramp, 120° angle at 14°C for 18 h. M: the λ PFG marker.

    Techniques Used: Agarose Gel Electrophoresis, Marker

    23) Product Images from "Genetic Characteristics of Borrelia coriaceae Isolates from the Soft Tick Ornithodoros coriaceus (Acari: Argasidae)"

    Article Title: Genetic Characteristics of Borrelia coriaceae Isolates from the Soft Tick Ornithodoros coriaceus (Acari: Argasidae)

    Journal: Journal of Clinical Microbiology

    doi:

    Southern hybridization of Apa I (a)-, Bam HI (b)-, and Hin dIII (c)-digested DNAs of Borrelia isolates with digoxigenin-labeled Co53 DNA as a probe. Lanes: 1, CA434; 2, CA435; 3, Co53; 4, B. parkeri ; 5, CA4. Southern hybridization and detection of DNA which hybridized to the probe were done as recommended by Boehringer Mannheim for the digoxigenin DNA labeling and detection kit.
    Figure Legend Snippet: Southern hybridization of Apa I (a)-, Bam HI (b)-, and Hin dIII (c)-digested DNAs of Borrelia isolates with digoxigenin-labeled Co53 DNA as a probe. Lanes: 1, CA434; 2, CA435; 3, Co53; 4, B. parkeri ; 5, CA4. Southern hybridization and detection of DNA which hybridized to the probe were done as recommended by Boehringer Mannheim for the digoxigenin DNA labeling and detection kit.

    Techniques Used: Hybridization, Labeling, DNA Labeling

    24) Product Images from "Identification and Characterization of Leuconostoc carnosum, Associated with Production and Spoilage of Vacuum-Packaged, Sliced, Cooked Ham"

    Article Title: Identification and Characterization of Leuconostoc carnosum, Associated with Production and Spoilage of Vacuum-Packaged, Sliced, Cooked Ham

    Journal: Applied and Environmental Microbiology

    doi:

    Cla I ribopatterns. Lanes 4 and 11, phage lambda DNA cleaved with Hin dIII as a fragment size marker; lane 1, Weissella viridescens ATCC 12706 T ; lane 2, Weissella halotolerans ATCC 35410 T ; lane 3, Weissella paramesenteroides DSM 20288 T ; lane 5, Leuconostoc mesenteroides subsp. mesenteroides DSM 20343 T ; lane 6, Leuconostoc mesenteroides subsp. cremoris CCUG 21965 T ; lane 7, Leuconostoc mesenteroides subsp. dextranicum DSM 20484 T ; lane 8, Leuconostoc pseudomesenteroides DSM 20193 T ; lane 9, Leuconostoc carnosum NCFB 2776 T ; lane 10, Leuconostoc gelidum NCFB 2775 T ; lane 12, Leuconostoc lactis CCUG 30064 T ; lane 13, Leuconostoc fallax CCUG 30061 T ; lane 14, Leuconostoc citreum ( Leuconostoc amelibiosum ) D1.
    Figure Legend Snippet: Cla I ribopatterns. Lanes 4 and 11, phage lambda DNA cleaved with Hin dIII as a fragment size marker; lane 1, Weissella viridescens ATCC 12706 T ; lane 2, Weissella halotolerans ATCC 35410 T ; lane 3, Weissella paramesenteroides DSM 20288 T ; lane 5, Leuconostoc mesenteroides subsp. mesenteroides DSM 20343 T ; lane 6, Leuconostoc mesenteroides subsp. cremoris CCUG 21965 T ; lane 7, Leuconostoc mesenteroides subsp. dextranicum DSM 20484 T ; lane 8, Leuconostoc pseudomesenteroides DSM 20193 T ; lane 9, Leuconostoc carnosum NCFB 2776 T ; lane 10, Leuconostoc gelidum NCFB 2775 T ; lane 12, Leuconostoc lactis CCUG 30064 T ; lane 13, Leuconostoc fallax CCUG 30061 T ; lane 14, Leuconostoc citreum ( Leuconostoc amelibiosum ) D1.

    Techniques Used: Lambda DNA Preparation, Marker

    25) Product Images from "Characterization of a Replication-Defective Human Immunodeficiency Virus Type 1 att Site Mutant That Is Blocked after the 3? Processing Step of Retroviral Integration"

    Article Title: Characterization of a Replication-Defective Human Immunodeficiency Virus Type 1 att Site Mutant That Is Blocked after the 3? Processing Step of Retroviral Integration

    Journal: Journal of Virology

    doi:

    Strategy for detecting 3′ processing activity in infected cell extracts. Hae III and Hin dIII cleave HIV-1 cDNA approximately 100 bp from the U3 and U5 ends, respectively. 3′ processing by integrase shortens the U3 minus strand and U5 plus strand by 2 nucleotides (nt). Following cell lysis and cleavage with Hae III and Hin dIII, both processed and nonprocessed strands were detected by indirect end labeling using strand-specific riboprobes.
    Figure Legend Snippet: Strategy for detecting 3′ processing activity in infected cell extracts. Hae III and Hin dIII cleave HIV-1 cDNA approximately 100 bp from the U3 and U5 ends, respectively. 3′ processing by integrase shortens the U3 minus strand and U5 plus strand by 2 nucleotides (nt). Following cell lysis and cleavage with Hae III and Hin dIII, both processed and nonprocessed strands were detected by indirect end labeling using strand-specific riboprobes.

    Techniques Used: Activity Assay, Infection, Lysis, End Labeling

    26) Product Images from "Proliferative Enterocolitis Associated with Dual Infection with Enteropathogenic Escherichia coli and Lawsonia intracellularis in Rabbits"

    Article Title: Proliferative Enterocolitis Associated with Dual Infection with Enteropathogenic Escherichia coli and Lawsonia intracellularis in Rabbits

    Journal: Journal of Clinical Microbiology

    doi:

    Southern blot of Hin dIII-digested genomic DNA from E. coli strains probed with the eaeA gene probe under conditions of high stringency. The lanes contain independent isolates from four rabbits with dual infections with EPEC and L. intracellularis , RDEC-1, and a nonpathogenic E. coli K-12 strain.
    Figure Legend Snippet: Southern blot of Hin dIII-digested genomic DNA from E. coli strains probed with the eaeA gene probe under conditions of high stringency. The lanes contain independent isolates from four rabbits with dual infections with EPEC and L. intracellularis , RDEC-1, and a nonpathogenic E. coli K-12 strain.

    Techniques Used: Southern Blot

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    Polymerase Chain Reaction:

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    Article Snippet: .. For floxCol12a1 screening, PCR products were digested with HindIII (NEB). .. in vitro Cre-recombination 200 ng of PCR products of cloned floxCol12a1 alleles from floxed mice or genomic Col12a1 PCR products of wildtype mice were incubated with Cre recombinase (NEB) and analyzed by 2% agarose gel electrophoresis according to the manufacture’s instruction.

    Article Title: Human DNA2 possesses a cryptic DNA unwinding activity that functionally integrates with BLM or WRN helicases
    Article Snippet: .. The PCR products were digested with BamHI and HindIII restriction endonucleases (New England Biolabs, Ipswich, MA) and ligated into a pFastBac1 vector (Invitrogen, Carlsbad, CA) generating pFB-His-hDNA2-FLAG. .. The D277A point mutation inactivating the hDNA2 nuclease was introduced with oligonucleotide pair 5'-GGCCTGAAGGGAAAGATCGCTGTCACAGTTGGAGTGAAG-3' and 5'-CTTCACTCCAACTGTGACAGCGATCTTTCCCTTCAGGCC-3' whereas the K654R point mutation abolishing the hDNA2 helicase was introduced with oligonucleotide pair 5'-GGCATGCCGGGAACTGGCAGGACAACCACTATCTGCACA-3' and 5'-TGTGCAGATAGTGGTTGTCCTGCCAGTTCCCGGCATGCC-3' using the QuikChange XL Site-directed mutagenesis kit (Agilent, Santa Clara, CA) according to manufacturer's recommendations.

    Transformation Assay:

    Article Title: Single-stranded DNA and RNA origami
    Article Snippet: .. In the next step, the two digested fragments were ligated with Eco RI and Hind III digested pGEM-7zf (−) vector, and transformed into E. coli cells (NEB stable competent cells). .. This was a three-fragment ligation to form a circular piece of DNA.

    other:

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    Agarose Gel Electrophoresis:

    Article Title: GES Extended-Spectrum ?-Lactamases in Acinetobacter baumannii Isolates in Belgium ▿
    Article Snippet: .. One microgram of natural plasmids was digested with 10 U of HindIII and 10 U of EcoRI (New England Biolabs, Hitchin, United Kingdom) at 37°C for 1 h. Restricted fragments were separated on a 0.8% agarose gel in 0.5× Tris-borate-EDTA (TBE) (44.5 mM Tris, 44.5 mM boric acid, 1 mM EDTA, pH 8.3) buffer containing 0.5 μg/ml of ethidium bromide for 1 h at 150 V and visualized under UV light. ..

    Article Title: Emergence of Resistance among USA300 Methicillin-Resistant Staphylococcus aureus Isolates Causing Invasive Disease in the United States ▿
    Article Snippet: .. HindIII (New England Biolabs, Beverly, MA)-restricted plasmid fragments were separated on a 0.75% agarose gel in Tris-borate-EDTA (TBE) buffer at 80 V for 5 h. The Trackit 1 Kb DNA ladder (Invitrogen, Carlsbad, CA) was used for sizing the linear fragments. .. Digoxigenin-labeled DNA probes were generated by PCR using the oligonucleotide primers shown in Table and plasmid DNA isolated from the following control organisms: S. aureus FPR3757 ( ) for tra E , tra I , rep A , and mup A , and S. aureus HIP11714 MI-1 (vancomycin resistant S. aureus [VRSA-1]) ( ) for aac 6 ′- aph 2 ″ and dfr A .

    Plasmid Preparation:

    Article Title: Single-stranded DNA and RNA origami
    Article Snippet: .. In the next step, the two digested fragments were ligated with Eco RI and Hind III digested pGEM-7zf (−) vector, and transformed into E. coli cells (NEB stable competent cells). .. This was a three-fragment ligation to form a circular piece of DNA.

    Article Title: Human DNA2 possesses a cryptic DNA unwinding activity that functionally integrates with BLM or WRN helicases
    Article Snippet: .. The PCR products were digested with BamHI and HindIII restriction endonucleases (New England Biolabs, Ipswich, MA) and ligated into a pFastBac1 vector (Invitrogen, Carlsbad, CA) generating pFB-His-hDNA2-FLAG. .. The D277A point mutation inactivating the hDNA2 nuclease was introduced with oligonucleotide pair 5'-GGCCTGAAGGGAAAGATCGCTGTCACAGTTGGAGTGAAG-3' and 5'-CTTCACTCCAACTGTGACAGCGATCTTTCCCTTCAGGCC-3' whereas the K654R point mutation abolishing the hDNA2 helicase was introduced with oligonucleotide pair 5'-GGCATGCCGGGAACTGGCAGGACAACCACTATCTGCACA-3' and 5'-TGTGCAGATAGTGGTTGTCCTGCCAGTTCCCGGCATGCC-3' using the QuikChange XL Site-directed mutagenesis kit (Agilent, Santa Clara, CA) according to manufacturer's recommendations.

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    Article Snippet: .. HindIII (New England Biolabs, Beverly, MA)-restricted plasmid fragments were separated on a 0.75% agarose gel in Tris-borate-EDTA (TBE) buffer at 80 V for 5 h. The Trackit 1 Kb DNA ladder (Invitrogen, Carlsbad, CA) was used for sizing the linear fragments. .. Digoxigenin-labeled DNA probes were generated by PCR using the oligonucleotide primers shown in Table and plasmid DNA isolated from the following control organisms: S. aureus FPR3757 ( ) for tra E , tra I , rep A , and mup A , and S. aureus HIP11714 MI-1 (vancomycin resistant S. aureus [VRSA-1]) ( ) for aac 6 ′- aph 2 ″ and dfr A .

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    New England Biolabs hin diii
    Southern blots of Hin <t>dIII</t> and Hin dIII-VDE digests of <t>DNA</t> from spo11 strains with inserts at HIS4 (top) and at URA3 (bottom). Gel labels are as in Figure 1 ; JM—joint molecule recombination intermediates. DOI: http://dx.doi.org/10.7554/eLife.19669.015
    Hin Diii, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 28 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    New England Biolabs hin diii digested genomic dna
    Southern blots of catalase-defective mutants. Shown are the Southern blots of Hin <t>dIII-digested</t> genomic <t>DNA</t> from the wild type, Y4Nal (lane 2), and catalase-negative mutants, Aa1393 (lane 3), Aa1394 (lane 4), and Aa1395 (lane 5). Lane 1 contains digoxigenin-labeled DNA molecular weight markers (shown in kilobases) (Boehringer Mannheim). Probes were digoxigenin-labeled DNA containing the kan gene (A) or the katA gene (B).
    Hin Diii Digested Genomic Dna, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 88/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    New England Biolabs hin diii restriction enzyme
    (a) Agarose gel (1%) electrophoresis of plasmids pCYLM01 and pCYLM02 obtained by gel elution. Lane M - λ DNA- Hin <t>dIII</t> and ϕ x174 DNA- Hae III digest Mix (Finnzymes); Lane 1 – Plasmid pCYLM01; Lane 2 – Plasmid pCYLM02; Lane 3 – Plasmids of Cylindrospermum <t>stagnale.</t> (b) Agarose gel (1%) electrophoresis of plasmids pCYLM01 and pCYLM02 digested with Hin dIII restriction enzyme. Lane 1 – Undigested plasmid pCYLM01; Lane 2 – pCYLM01 plasmid digested with Hin dIII; Lane 3 – Undigested plasmid pCYLM02; Lane 4 – pCYLM02 plasmid digested with Hin dIII; Lane 5 – Total plasmid of Cylindrospermum stagnale.
    Hin Diii Restriction Enzyme, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 93/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Southern blots of Hin dIII and Hin dIII-VDE digests of DNA from spo11 strains with inserts at HIS4 (top) and at URA3 (bottom). Gel labels are as in Figure 1 ; JM—joint molecule recombination intermediates. DOI: http://dx.doi.org/10.7554/eLife.19669.015

    Journal: eLife

    Article Title: Local chromosome context is a major determinant of crossover pathway biochemistry during budding yeast meiosis

    doi: 10.7554/eLife.19669

    Figure Lengend Snippet: Southern blots of Hin dIII and Hin dIII-VDE digests of DNA from spo11 strains with inserts at HIS4 (top) and at URA3 (bottom). Gel labels are as in Figure 1 ; JM—joint molecule recombination intermediates. DOI: http://dx.doi.org/10.7554/eLife.19669.015

    Article Snippet: Recombination products were detected on Southern blots containing genomic DNA digested with Hin dIII and VDE (P I-Sce I, New England Biolabs), using specific buffer for P I-Sce I.

    Techniques:

    Spo11-initiated events at the two insert loci. ( A ) Spo11-catalyzed DSBs are more frequent at HIS4 that at URA3 . Left—Southern blots of Eco RI digests of DNA from vde∆ strains, probed with pBR322 sequences, showing Spo11-DSBs in the Parent 2 insert (see Figure 1 ) in resection/repair-deficient sae2∆ mutant strains. Right—location of DSBs and probe and DSB frequencies (average of 7 and 8 hr samples from a single experiment; error bars represent range). Spo11-DSBs in the Parent 1 inserts at HIS4 and URA3 were at different locations within the insert, but displayed similar ratios between the two loci (data not shown). ( B ) Southern blots of Hin dIII digests of DNA from vde∆ strains, to detect total Spo11-initiated crossovers. ( C ) Southern blots of Hin dIII-VDE double digests of the same samples, to determine the background contribution of Spo11-initiated COs in subsequent experiments measuring VDE-initiated COs, which will be VDE-resistant due to conversion of the VRS site to VRS103 . Probes were as shown in Figure 1 . ( D ) Quantification of data in panels B (total COs; filled circles) and C (VDE-resistant COs; open circles). Data are from a single experiment. DOI: http://dx.doi.org/10.7554/eLife.19669.004

    Journal: eLife

    Article Title: Local chromosome context is a major determinant of crossover pathway biochemistry during budding yeast meiosis

    doi: 10.7554/eLife.19669

    Figure Lengend Snippet: Spo11-initiated events at the two insert loci. ( A ) Spo11-catalyzed DSBs are more frequent at HIS4 that at URA3 . Left—Southern blots of Eco RI digests of DNA from vde∆ strains, probed with pBR322 sequences, showing Spo11-DSBs in the Parent 2 insert (see Figure 1 ) in resection/repair-deficient sae2∆ mutant strains. Right—location of DSBs and probe and DSB frequencies (average of 7 and 8 hr samples from a single experiment; error bars represent range). Spo11-DSBs in the Parent 1 inserts at HIS4 and URA3 were at different locations within the insert, but displayed similar ratios between the two loci (data not shown). ( B ) Southern blots of Hin dIII digests of DNA from vde∆ strains, to detect total Spo11-initiated crossovers. ( C ) Southern blots of Hin dIII-VDE double digests of the same samples, to determine the background contribution of Spo11-initiated COs in subsequent experiments measuring VDE-initiated COs, which will be VDE-resistant due to conversion of the VRS site to VRS103 . Probes were as shown in Figure 1 . ( D ) Quantification of data in panels B (total COs; filled circles) and C (VDE-resistant COs; open circles). Data are from a single experiment. DOI: http://dx.doi.org/10.7554/eLife.19669.004

    Article Snippet: Recombination products were detected on Southern blots containing genomic DNA digested with Hin dIII and VDE (P I-Sce I, New England Biolabs), using specific buffer for P I-Sce I.

    Techniques: Mutagenesis

    70–80% of VDE-DSBs are repaired. ( A ) Fraction of inserts remaining, calculated using Hin dIII digests (see Figure 1 ). For the arg4-VRS103 insert, the ratio (Parent 2 + CO2)/ (0.5 x LC) was calculated at 9 hr, and was then normalized to the 0 hr value. For the arg4-VRS insert, a similar calculation was made: (Parent 1 + NCO + CO1)/(0.5 x LC) ( B ) Relative recovery of interhomolog recombination products, calculated using Hin dIII-VDE double digests (see Figure 1 ). The sum of CO (average of CO1 and CO2) and NCO frequencies was divided by the frequency of total DSBs, as calculated in Figure 2A . Data are the average of two independent experiments; error bars represent range. DOI: http://dx.doi.org/10.7554/eLife.19669.006

    Journal: eLife

    Article Title: Local chromosome context is a major determinant of crossover pathway biochemistry during budding yeast meiosis

    doi: 10.7554/eLife.19669

    Figure Lengend Snippet: 70–80% of VDE-DSBs are repaired. ( A ) Fraction of inserts remaining, calculated using Hin dIII digests (see Figure 1 ). For the arg4-VRS103 insert, the ratio (Parent 2 + CO2)/ (0.5 x LC) was calculated at 9 hr, and was then normalized to the 0 hr value. For the arg4-VRS insert, a similar calculation was made: (Parent 1 + NCO + CO1)/(0.5 x LC) ( B ) Relative recovery of interhomolog recombination products, calculated using Hin dIII-VDE double digests (see Figure 1 ). The sum of CO (average of CO1 and CO2) and NCO frequencies was divided by the frequency of total DSBs, as calculated in Figure 2A . Data are the average of two independent experiments; error bars represent range. DOI: http://dx.doi.org/10.7554/eLife.19669.006

    Article Snippet: Recombination products were detected on Southern blots containing genomic DNA digested with Hin dIII and VDE (P I-Sce I, New England Biolabs), using specific buffer for P I-Sce I.

    Techniques:

    Southern blots of Hin dIII and Hin dIII-VDE digests of DNA from HIS4 insert-containing strains (top) and from URA3 insert-contaning strains (bottom). Probes and gel labels are as in Figure 1 ; JM—joint molecule recombination intermediates. DOI: http://dx.doi.org/10.7554/eLife.19669.009

    Journal: eLife

    Article Title: Local chromosome context is a major determinant of crossover pathway biochemistry during budding yeast meiosis

    doi: 10.7554/eLife.19669

    Figure Lengend Snippet: Southern blots of Hin dIII and Hin dIII-VDE digests of DNA from HIS4 insert-containing strains (top) and from URA3 insert-contaning strains (bottom). Probes and gel labels are as in Figure 1 ; JM—joint molecule recombination intermediates. DOI: http://dx.doi.org/10.7554/eLife.19669.009

    Article Snippet: Recombination products were detected on Southern blots containing genomic DNA digested with Hin dIII and VDE (P I-Sce I, New England Biolabs), using specific buffer for P I-Sce I.

    Techniques:

    Southern blots of Hin dIII and Hin dIII-VDE digests of DNA from HIS4 insert-containing strains (top) and from URA3 insert-contaning strains (bottom). Gel labels are as in Figure 1 ; JM—joint molecule recombination intermediates. In the gel with Hin DIII digests of samples from a pch2∆ mm4-mn yen1∆ slx1∆ strain with inserts at URA3 , the 9 hr sample was originally loaded between the 4 and 5 hr samples; this image was cut and spliced as indicated by vertical lines for presentation purposes. DOI: http://dx.doi.org/10.7554/eLife.19669.012

    Journal: eLife

    Article Title: Local chromosome context is a major determinant of crossover pathway biochemistry during budding yeast meiosis

    doi: 10.7554/eLife.19669

    Figure Lengend Snippet: Southern blots of Hin dIII and Hin dIII-VDE digests of DNA from HIS4 insert-containing strains (top) and from URA3 insert-contaning strains (bottom). Gel labels are as in Figure 1 ; JM—joint molecule recombination intermediates. In the gel with Hin DIII digests of samples from a pch2∆ mm4-mn yen1∆ slx1∆ strain with inserts at URA3 , the 9 hr sample was originally loaded between the 4 and 5 hr samples; this image was cut and spliced as indicated by vertical lines for presentation purposes. DOI: http://dx.doi.org/10.7554/eLife.19669.012

    Article Snippet: Recombination products were detected on Southern blots containing genomic DNA digested with Hin dIII and VDE (P I-Sce I, New England Biolabs), using specific buffer for P I-Sce I.

    Techniques:

    Mapping and tandem arrangement of DXZ4 in primates . (a) Direct-labeled fluorescence in situ hybridization (FISH) of human DXZ4 BAC clone (2272M5; red) and human PLS3 BAC clone (268A15; green) to male and female rhesus macaque metaphase chromosomes. White arrows point to the hybridizing X chromosome. Metaphase chromosomes were counterstained with DAPI, and converted to gray-scale to assist in visualizing the FISH signals. (b) Southern blot of Xba I digested primate genomic DNA separated by pulsed field gel electrophoresis, hybridized with a human digoxigenin-labeled DXZ4 probe. Primates and group are listed along the top and gender indicated by M (male) or F (female), including rhesus macaque (R. Macaque), pig-tailed macaque (P-T. Macaque), common squirrel monkey (Sq. Monkey) and black-handed spider monkey (Sp. Monkey). Size in kilobases is given to the left. (c) Ethidium bromide stained 0.9% agarose gel showing green monkey and macaque BAC DNA digested with the restriction endonuclease Hin dIII and separated by gel electrophoresis. The sizes of the molecular weight marker are given to the left in kilobases.

    Journal: Genome Biology

    Article Title: Characterization of DXZ4 conservation in primates implies important functional roles for CTCF binding, array expression and tandem repeat organization on the X chromosome

    doi: 10.1186/gb-2011-12-4-r37

    Figure Lengend Snippet: Mapping and tandem arrangement of DXZ4 in primates . (a) Direct-labeled fluorescence in situ hybridization (FISH) of human DXZ4 BAC clone (2272M5; red) and human PLS3 BAC clone (268A15; green) to male and female rhesus macaque metaphase chromosomes. White arrows point to the hybridizing X chromosome. Metaphase chromosomes were counterstained with DAPI, and converted to gray-scale to assist in visualizing the FISH signals. (b) Southern blot of Xba I digested primate genomic DNA separated by pulsed field gel electrophoresis, hybridized with a human digoxigenin-labeled DXZ4 probe. Primates and group are listed along the top and gender indicated by M (male) or F (female), including rhesus macaque (R. Macaque), pig-tailed macaque (P-T. Macaque), common squirrel monkey (Sq. Monkey) and black-handed spider monkey (Sp. Monkey). Size in kilobases is given to the left. (c) Ethidium bromide stained 0.9% agarose gel showing green monkey and macaque BAC DNA digested with the restriction endonuclease Hin dIII and separated by gel electrophoresis. The sizes of the molecular weight marker are given to the left in kilobases.

    Article Snippet: Individual DXZ4 monomers from BAC clones CH250-131A6 and CH252-338G16 were generated by first performing a Hin dIII digestion on the BAC clone DNA, gel purifying the 3-kb fragment and cloning into calf intestinal alkaline phosphatase (NEB) treated Hin dIII cut pBluescript-II (Agilent Technologies, Santa Clara, CA, USA).

    Techniques: Labeling, Fluorescence, In Situ Hybridization, Fluorescence In Situ Hybridization, BAC Assay, Southern Blot, Pulsed-Field Gel, Electrophoresis, Staining, Agarose Gel Electrophoresis, Nucleic Acid Electrophoresis, Molecular Weight, Marker

    Southern blots of catalase-defective mutants. Shown are the Southern blots of Hin dIII-digested genomic DNA from the wild type, Y4Nal (lane 2), and catalase-negative mutants, Aa1393 (lane 3), Aa1394 (lane 4), and Aa1395 (lane 5). Lane 1 contains digoxigenin-labeled DNA molecular weight markers (shown in kilobases) (Boehringer Mannheim). Probes were digoxigenin-labeled DNA containing the kan gene (A) or the katA gene (B).

    Journal: Journal of Bacteriology

    Article Title: Direct Selection of IS903 Transposon Insertions by Use of a Broad-Host-Range Vector: Isolation of Catalase-Deficient Mutants of Actinobacillus actinomycetemcomitans

    doi:

    Figure Lengend Snippet: Southern blots of catalase-defective mutants. Shown are the Southern blots of Hin dIII-digested genomic DNA from the wild type, Y4Nal (lane 2), and catalase-negative mutants, Aa1393 (lane 3), Aa1394 (lane 4), and Aa1395 (lane 5). Lane 1 contains digoxigenin-labeled DNA molecular weight markers (shown in kilobases) (Boehringer Mannheim). Probes were digoxigenin-labeled DNA containing the kan gene (A) or the katA gene (B).

    Article Snippet: A 1-μg portion of Hin dIII-digested genomic DNA was ligated with 10 U of DNA ligase (New England Biolabs) in a 50-μl reaction mixture at 14°C for 16 h. The large reaction volume was used to facilitate intramolecular ligation.

    Techniques: Labeling, Molecular Weight

    (a) Agarose gel (1%) electrophoresis of plasmids pCYLM01 and pCYLM02 obtained by gel elution. Lane M - λ DNA- Hin dIII and ϕ x174 DNA- Hae III digest Mix (Finnzymes); Lane 1 – Plasmid pCYLM01; Lane 2 – Plasmid pCYLM02; Lane 3 – Plasmids of Cylindrospermum stagnale. (b) Agarose gel (1%) electrophoresis of plasmids pCYLM01 and pCYLM02 digested with Hin dIII restriction enzyme. Lane 1 – Undigested plasmid pCYLM01; Lane 2 – pCYLM01 plasmid digested with Hin dIII; Lane 3 – Undigested plasmid pCYLM02; Lane 4 – pCYLM02 plasmid digested with Hin dIII; Lane 5 – Total plasmid of Cylindrospermum stagnale.

    Journal: Saudi Journal of Biological Sciences

    Article Title: Isolation and characterization of two novel plasmids pCYM01 and pCYM02 of Cylindrospermum stagnale

    doi: 10.1016/j.sjbs.2019.11.017

    Figure Lengend Snippet: (a) Agarose gel (1%) electrophoresis of plasmids pCYLM01 and pCYLM02 obtained by gel elution. Lane M - λ DNA- Hin dIII and ϕ x174 DNA- Hae III digest Mix (Finnzymes); Lane 1 – Plasmid pCYLM01; Lane 2 – Plasmid pCYLM02; Lane 3 – Plasmids of Cylindrospermum stagnale. (b) Agarose gel (1%) electrophoresis of plasmids pCYLM01 and pCYLM02 digested with Hin dIII restriction enzyme. Lane 1 – Undigested plasmid pCYLM01; Lane 2 – pCYLM01 plasmid digested with Hin dIII; Lane 3 – Undigested plasmid pCYLM02; Lane 4 – pCYLM02 plasmid digested with Hin dIII; Lane 5 – Total plasmid of Cylindrospermum stagnale.

    Article Snippet: 2.6 Cloning of C. stagnale plasmids The vector pBlueScriptSK(-) was digested with Hin dIII restriction enzyme, dephosphorylated with Calf Intestinal Alkaline Phosphatase (NEB) and purified by phenol: chloroform (1:1; v/v) extraction method.

    Techniques: Agarose Gel Electrophoresis, Electrophoresis, Plasmid Preparation

    (a) Agarose gel (0.8%) electrophoresis of total DNA extracted from Cylindrospermum stagnale . Arrows indicate the extra chromosomal DNA. M – λ DNA- Hin dIII digest and ϕ x174 DNA- Hae III digest Mix (Finnzymes); Lane 1 – Total DNA of Cylindrospermum stagnale . (b) Agarose gel (0.8%) electrophoresis of plasmid DNA isolated from Cylindrospermum stagnale. Lane M – λ DNA- Hin dIII digest and ϕ x174 DNA- Hae III digest Mix (Finnzymes) ; Lanes 1 to 4 – Plasmid profile of Cylindrospermum stagnale isolated from different batches of cultures.

    Journal: Saudi Journal of Biological Sciences

    Article Title: Isolation and characterization of two novel plasmids pCYM01 and pCYM02 of Cylindrospermum stagnale

    doi: 10.1016/j.sjbs.2019.11.017

    Figure Lengend Snippet: (a) Agarose gel (0.8%) electrophoresis of total DNA extracted from Cylindrospermum stagnale . Arrows indicate the extra chromosomal DNA. M – λ DNA- Hin dIII digest and ϕ x174 DNA- Hae III digest Mix (Finnzymes); Lane 1 – Total DNA of Cylindrospermum stagnale . (b) Agarose gel (0.8%) electrophoresis of plasmid DNA isolated from Cylindrospermum stagnale. Lane M – λ DNA- Hin dIII digest and ϕ x174 DNA- Hae III digest Mix (Finnzymes) ; Lanes 1 to 4 – Plasmid profile of Cylindrospermum stagnale isolated from different batches of cultures.

    Article Snippet: 2.6 Cloning of C. stagnale plasmids The vector pBlueScriptSK(-) was digested with Hin dIII restriction enzyme, dephosphorylated with Calf Intestinal Alkaline Phosphatase (NEB) and purified by phenol: chloroform (1:1; v/v) extraction method.

    Techniques: Agarose Gel Electrophoresis, Electrophoresis, Plasmid Preparation, Isolation

    Agarose gel (1%) electrophoresis of heat-treated and unheated plasmid DNAs from Cylindrospermum stagnale studied together with heat-treated and unheated bacterial plasmids pUC18 and pBR322 and linear λ DNA. Heat-treated samples of DNA were obtained by heating the samples in the presence of 0.1% (w/v) sarkosyl for 2 min at 100 °C followed by quick cooling. The arrows indicate Covalently Closed Circular (CCC) DNA. Lane M - λ DNA- Hin dIII and ϕ x174 DNA- Hae III digest Mix (Finnzymes); Lane 1 – Unheated Cylindrospermum stagnale plasmids ; Lane 2 – Heat-treated Cylindrospermum stagnale plasmids; Lane 3 – Unheated pUC18 plasmid; Lane 4 – Heat-treated pUC18 plasmid; Lane 5 – pBR322 plasmid; Lane 6 – Heat treated pBR322 plasmid; Lane 7 – Unheated λ DNA; Lane 8 – Heat treated λ DNA.

    Journal: Saudi Journal of Biological Sciences

    Article Title: Isolation and characterization of two novel plasmids pCYM01 and pCYM02 of Cylindrospermum stagnale

    doi: 10.1016/j.sjbs.2019.11.017

    Figure Lengend Snippet: Agarose gel (1%) electrophoresis of heat-treated and unheated plasmid DNAs from Cylindrospermum stagnale studied together with heat-treated and unheated bacterial plasmids pUC18 and pBR322 and linear λ DNA. Heat-treated samples of DNA were obtained by heating the samples in the presence of 0.1% (w/v) sarkosyl for 2 min at 100 °C followed by quick cooling. The arrows indicate Covalently Closed Circular (CCC) DNA. Lane M - λ DNA- Hin dIII and ϕ x174 DNA- Hae III digest Mix (Finnzymes); Lane 1 – Unheated Cylindrospermum stagnale plasmids ; Lane 2 – Heat-treated Cylindrospermum stagnale plasmids; Lane 3 – Unheated pUC18 plasmid; Lane 4 – Heat-treated pUC18 plasmid; Lane 5 – pBR322 plasmid; Lane 6 – Heat treated pBR322 plasmid; Lane 7 – Unheated λ DNA; Lane 8 – Heat treated λ DNA.

    Article Snippet: 2.6 Cloning of C. stagnale plasmids The vector pBlueScriptSK(-) was digested with Hin dIII restriction enzyme, dephosphorylated with Calf Intestinal Alkaline Phosphatase (NEB) and purified by phenol: chloroform (1:1; v/v) extraction method.

    Techniques: Agarose Gel Electrophoresis, Electrophoresis, Plasmid Preparation, Countercurrent Chromatography